Two new triterpenoid glycosides from Curculigo orchioides

Article abstract of DOI:10.1080/10286020.2012.656607

Two new cycloartane triterpenoid glycosides, named curculigosaponin N and curculigosaponin O, were isolated from rhizomes of Curculigo orchioides Gaertn. Their structures were elucidated on the basis of comprehensive spectroscopic analysis including IR, M

Full text of DOI:10.1080/10286020.2012.656607

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Two new triterpenoid glycosides from
Curculigo orchioides
Ai-Xue Zuo a b , Yong Shen a , Zhi-Yong Jiang a , Xue-Mei Zhang a ,
Jun Zhou a , Jun Lü c & Ji-Jun Chen a
a State Key Laboratory of Phytochemistry and Plant Resources
in West China, Kunming Institute of Botany, Chinese Academy of
Sciences, Kunming, 650204, China
b Yunnan University of Traditional Chinese Medicine, Kunming,
650500, China
c Kunming Jingbiao Biosciences R&D Co. Ltd, Kunming, 650000,
China
Available online: 09 Feb 2012
To cite this article: Ai-Xue Zuo, Yong Shen, Zhi-Yong Jiang, Xue-Mei Zhang, Jun Zhou, Jun Lü &
Ji-Jun Chen (2012): Two new triterpenoid glycosides from Curculigo orchioides , Journal of Asian
Natural Products Research, 14:5, 407-412
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Journal of Asian Natural Products Research
Vol. 14, No. 5, May 2012, 407–412
Two new triterpenoid glycosides from Curculigo orchioides
Ai-Xue Zuoab, Yong Shena, Zhi-Yong Jianga, Xue-Mei Zhanga, Jun Zhoua, Jun Lu¨c and
Ji-Jun Chena*
aState Key Laboratory of Phytochemistry and Plant Resources in West China, Kunming Institute of
Botany, Chinese Academy of Sciences, Kunming 650204, China; bYunnan University of Traditional
Chinese Medicine, Kunming 650500, China; Kunming Jingbiao Biosciences R&D Co. Ltd,
c
Kunming 650000, China
(Received 30 December 2011; final version received 9 January 2012)
Two new cycloartane triterpenoid glycosides, named curculigosaponin N and
curculigosaponin O, were isolated from rhizomes of Curculigo orchioides Gaertn.
Their structures were elucidated on the basis of comprehensive spectroscopic analysis
including IR, MS, 1D, and 2D NMR (HSQC and HMBC).
Keywords: Curculigo orchioides; cycloartane triterpenoid glycoside; curculigosapo-
nins NZO
1. Introduction
find structurally new chemical constituents
from this medicinal plant, we explored its
phytochemical composition. Our reinvesti-
gation resulted in the isolation of two new
cycloartane triterpenoid glycosides, named
curculigosaponin N and curculigosaponin
O. This paper deals with the isolation and
structural elucidation of the two new
compounds (1 and 2; Figure 1).
Curculigo orchioides Gaertn., belonged to
the Hypoxidaceae family and named ‘Xian-
Mao’ in Pharmacopoeia of China, mainly
distributed in Japan, China, Malaysia, India,
and Australia [1]. Its rhizomes have been
employed as an analeptic agent for the
treatment of decline in strength, against
jaundice and asthma [1]. Previous investi-
gation on this plant revealed that it
contained cycloartane triterpenoid saponins
[2–5], phenolic glycosides [6–8], and
chlorophenyl glycosides [9]. Some com-
pounds from C. orchioides exhibited
biological activities including stimulation
of the immune response [10,11], antioxida-
tive activities [8], antidepression activities
[12], and cytotoxic activities [4,5].
Recently, several novel phenolic glycosides
[13–16] in this genus have been elucidated
by our group, meanwhile, some isolated
phenolic derivatives displayed significant
antidepressant effect [12] and anti-hepatitis
B virus (HBV) activities [13]. To further
2. Results and discussion
Compound 1 was isolated as an amorphous
powder with ½aꢀ1d5:3 2 3:41 (c 0.18,
MeOH); its molecular formula was deter-
mined to be C42H72O14 based on HR-ESI-
MS at m/z 799.4841 [M 2 H]2. Negative
mode FAB-MS exhibited quasi-molecular
ion and fragment ions at m/z 799 [M 2 H]2,
653
[M 2 H 2 146]2,
and
491
[M 2 H 2 146 2 162]2, suggesting the
presence of hexose moiety in the molecule
of compound 1. Acid hydrolysis of com-
pound 1 with 2 M H2SO4 liberated rham-
nose and glucose detected by comparison
*Corresponding author. Email: chenjj@mail.kib.ac.cn
ISSN 1028-6020 print/ISSN 1477-2213 online
q 2012 Taylor & Francis
http://dx.doi.org/10.1080/10286020.2012.656607
http://www.tandfonline.com

408
A.-X. Zuo et al.
group. This inference was further supported
OH
21
18
27
by the correlations observed from H-26 and
H-27 to C-25 in HMBC experiment. In
addition, the HMBC correlations observed
from H-10 at dH 4.93 (1H, d, J ¼ 6.9 Hz) to
C-3 and fromH-100 atdH 6.58(1H, br.s) toC-
20 demonstrated that the inner glucopyr-
anosyl unit was attached to C-3 of aglycone,
and the rhamnopyranosylmoietywas linked
to C-20 of the inner glucose (Figure 2).
Based on the above evidence, compound 1
was elucidated as 25-hydroxyl-curculigosa-
ponin L, and named curculigosaponin N
(1; Figure 1).
23
25
OH
11
HO
OH
17
13
19
1
9
15
10
3
30
7
5
RO
28
29
Curculigosaponin N (1)
R = Rha(1-2)Glc
Curculigosaponin O (2) R = Glc(1-3)Glc(1-2)Glc
Figure 1. The structures of compounds 1
and 2.
Compound 2, obtained as an amor-
17:8
with authentic samples on paper chroma-
tography (PC). IR showed the absorption
bands for hydroxy (3438 cm21), saturated
methenes (2926 cm21), and glycosidic
phous powder with ½aꢀd þ 4:62 (c 0.15,
MeOH), had the molecular formula
C48H82O20 as established from the posi-
tive-mode HR-ESI-MS at m/z 1001.5290
[M þ Na]þ. Negative-mode FAB-MS
spectrum showed quasi-molecular ion and
fragment ions at m/z 977 [M 2 H]2, 815
[M 2 162]2, 653 [M 2 162 2 162]2, and
491[M 2 162 2 162 2 162]2, suggesting
the presence of three hexoses in the
molecule of compound 2. Hydrolysis of 2
with 2 M H2SO4 in MeOH revealed glucose
as the only sugar moiety identified by
comparison with authentic sample on PC.
IR spectrum indicated the presence of
hydroxyl (3423 cm21) and glycosidic
1
bonds (1036 cm21). H NMR spectrum of
compound 1 revealed the presence of
characteristic cyclopropane methylene pro-
ton resonances at dH 0.26 (1H, d,
J ¼ 3.2 Hz, H-19a) and 0.43 (1H, d,
J ¼ 3.2 Hz, H-19b), six singlet methyl
resonances at dH 1.22, 1.26, 1.28, 1.30,
1.10, and 1.43 (each 3H, s, H-18, H-28, H-
29, H-30, H-25, H-26) assignable to
cycloartane-type aglycone, together with
two anomeric proton signals at dH 4.93 (1H,
d, J ¼ 6.9 Hz, H-10) and 6.58 (1H, br. s, H-
100), implying the presence of one b-linked
glucosyl moiety and one a-linked rhamno-
syl moiety. Rhamnosyl unit further sup-
ported by the methyl signal at d 1.46 (3H, d,
J ¼ 6.1Hz, H-600). The 13C NMR spectrum
of compound 1 exhibited 42 carbon signals,
of which aglycone included seven methyl
carbon signals at dC 15.7, 17.4, 18.8, 22.0,
25.7, 25.8, and26.5andfiveoxygen-bearing
carbons at dC 72.1, 72.5, 72.6, 77.1, and
88.6. The 13C NMR data also confirmed the
presence of two sugar moieties with
anomeric carbons at dC 105.4 (d) and
101.8 (d), respectively. Above NMR data
were similar to those of curculigosaponin L
[2] except that C-25 in compound 1 was
shifted downfield to dC 72.6, indicating that
C-25 was substituted by a hydroxyl
1
(1077 cm21) groups. The H NMR spec-
trum displayed characteristic of the C-19
methylene protons of cyclopropane ring for
a cycloartane triterpene at dH 0.47 (1H, d,
J ¼ 3.6 Hz, H-19a) and 0.56 (1H, d,
J ¼ 3.6 Hz, H-19b), seven methyl signals
at dH 0.91, 1.00, 1.13, 1.17, 1.19 (each 3H,
s, H-29, H-18, H-28, H-26, H-27), and
1.08–1.09 (6H, overlap, H-21, H-30)
attributable to cycloartane-type aglycone,
as well as three anomeric proton signals at
dH 4.47 (1H, d, J ¼ 7.0 Hz, H-10), 4.58 (1H,
d, J ¼ 7.8 Hz, H-100), and 4.76 (1H, d,
J ¼ 7.7 Hz, H-10) assigned to three b-
linked sugar moieties. 13C NMR spectrum
displayed 48 signals, of which the anomeric
carbons of three sugar moieties at dC 105.2
(d), 105.2 (d), and 104.2 (d) were observed.

Journal of Asian Natural Products Research
409
OH
21
18
23
CH3
OH
11
HO
OH CH3
OH
17
19
13
21
18
1
9
23
CH3
OH
15
10
28
3
11
HO
19
OR2 CH3
7
17
5
13
O
HO
1
9
30
1′
29
O
15
10
OH
3
30
7
5
H
HO
O
OH
O
29
28
HO
O
OH
1′
O
O
1″
H
OH
OH
O
H
OH
OH
O
HO
1″
CH3
O
H
OH
1″′
H
OH
OH
OH
OH
Figure 2. The key HMBC correlations of compounds 1 and 2.
Comparing the NMR data of compound 2
with those of curculigosaponin K [2]
indicated that they were analogous in
structure except that C-25 was shifted
downfield from dC 31.1 (d) to dC 73.8 (s),
suggesting a hydroxyl linked to C-25; to
further substantiate the location of the
hydroxy group, HMBC experiment was
conducted, in which the correlations from
H-26 at dH 1.17 (3H, s) and H-27 at dH 1.19
(3H, s) to C-25 at dC 73.8 (s) were observed.
Therefore, compound 2 was determined as
standard; chemical shifts (d) were expressed
in ppm and coupling constants (J) in Hz.
FAB-MS was recorded on a VG Autospec
3000 mass spectrometer (Manchester,
England); ESI and HR-ESI-MS were
taken on an API Qstar-Pulsar-1 mass
spectrometer (Applied Biosystems/MDS
Sciex, Vaughan, ON, Canada). Column
chromatography (CC) separations were
performed on silica gel (200–300 mesh,
Qingdao Meigao Chemical Co., Ltd.,
Qingdao, China), Al2O3 (Shanghai Wusi
Chemical Reagents Company, Shanghai,
China), D101 macroporous resins (Tianjin
Pesticide Chemical Company, Tianjin,
China), Sephadex LH-20 (Pharmacia Fine
Chemical Co. Ltd., Uppsala, Sweden), and
LiChroprep RP-18 (40–63 mm; Merck,
Darmstadt, Germany). Fractions were mon-
itored by thin-layer chromatography and
visualized by spraying with 10% H2SO4 in
EtOH followed by heating.
25-hydroxyl-curculigosaponin
named curculigosaponin O (Figure 2).
K and
3. Experimental
3.1 General experimental procedures
Optical rotations were performed on a
Horiba SEPA-300 polarimeter (Tokyo,
Japan). IR spectra were recorded on a Bio-
Rad FTS-135 spectrometer (Richmond,
VA, USA) with KBr pellets, n in cm21
.
UV spectra were measured on a UV-210A
spectrometer (Shimadzu, Japan); NMR
spectra were conducted on Bruker AV-400
or DRX-500 spectrometer (Karlsruhe,
Germany) with tetramethylsilane as internal
3.2 Plant material
The rhizomes of C. orchioides Gaertn.
were collected in Wenshan county, Yun-
nan Province, China, in November 2005,

410
A.-X. Zuo et al.
and authenticated by Prof. Dr Li-Gong
Lei, Kunming Institute of Botany, Chinese
Academy of Sciences. A voucher speci-
men (No. 20051106) has been deposited in
the Group of Anti-virus and Natural
Medicinal Chemistry, Kunming Institute
of Botany, Chinese Academy of Sciences.
3438, 2926, 1629, 1429, 1036, 876, 537,
481 cm21; 1H NMR (400 MHz, C5D5N) d:
0.26 (1H, d, J ¼ 3.2 Hz, H-19a), 0.43 (1H,
d, J ¼ 3.2 Hz, H-19b), 1.22, 1.26, 1.28,
1.30, 1.10, 1.43 (each 3H, s, H-18, H-28,
H-29, H-30, H-25, H-26), 1.28–1.30
(3H, overlap, H-21), 4.93 (1H, d,
J ¼ 6.9 Hz, H-10), 6.58 (1H, br. s, H-100),
1.46 (3H, d, J ¼ 6.1 Hz, H-600), 3.41
(1H, dd, J ¼ 11.2, 3.6 Hz, H-3), 3.95–
3.97(1H, m, H-11), 3.90–3.91 (1H, m, H-
16), 4.26–4.29 (1H, m, H-24); for
13C NMR (100 MHz, C5D5N) spectral
data, see Table 1. (2) FAB-MS: m/z 799
[M 2 H]2, 653 [M 2 H 2 146]2, 491
[M 2 H 2 146 2 162]2; (2) HR-ESI-
MS: m/z 799.4841 [M 2 H]2 (cald for
C42H71O14, 799.4843).
3.3 Extraction and isolation
The air-dried, powdered rhizomes of
C. orchioides (200 kg) were extracted
with 70% EtOH (each 1000 liters, 2 h)
three times under reflux to yield an extract
which was combined and concentrated to a
small volume (600 liters) and submitted to
CC (macroporous resin D101, 200 kg),
with gradient elution of H2O, 10% EtOH–
H2O, 40% EtOH–H2O, 70% EtOH–H2O,
and 90% EtOH–H2O to afford five
fractions (1–5). The fraction 4 (70%
EtOH–H2O eluted, 200 g) was subjected
to Al2O3 CC subsequently eluted with
3.3.2 Curculigosaponin O (2)
17:8
Amorphous powder; C48H82O20; ½aꢀD
þ
4:62 (c 0.15, MeOH); UV (MeOH) lmax
(log e): 202 (3.73) nm; IR (KBr) nmax:
3423, 2936, 1641, 1457, 1384, 1077,
635 cm21; H NMR (400 MHz, CD3OD)
EtOAc–EtOH–H2O
(8:2:0.2)
and
EtOAc–EtOH–H2O (7:3:0.5) to afford
fractions A and B.
1
Fraction A (60.0 g) was successively
subjected to RP-18 CC eluted with
MeOH–H2O (4:6–7:3) to afford fractions
A1–3. Fraction A2 (22.0 g) was performed
on silica gel CC eluted with CHCl3–
MeOH–H2O (8:2:0.3) to give a residue
(2.0 g) that was submitted to Sephadex
LH-20 CC developed with CHCl3–MeOH
(1:1) and further purified by silica gel CC
with EtOAc–EtOH–H2O (8:2:0.3) as
solvent to yield compound 2 (290 mg).
Fraction A3 (9.0 g) was applied to a silica
gel CC eluted with CHCl3–MeOH–H2O
(8.5:1.5:0.2) to give four portions. The
fourth portion (1.3 g) was purified on RP-
18 CC eluted with MeOH–H2O (5:5) to
afford compound 1 (221 mg).
d: 0.47 (1H, d, J ¼ 3.6 Hz, H-19a), 0.56
(1H, d, J ¼ 3.6 Hz, H-19b), 0.91, 1.00,
1.13, 1.17, 1.19 (each 3H, s, H-29, H-18,
H-28, H-26, H-27), 1.08–1.09 (6H, over-
lap, H-21, H-30), 4.47 (1H, d, J ¼ 7.0 Hz,
H-10), 4.58 (1H, d, J ¼ 7.8 Hz, H-100),
4.76 (1H, d, J ¼ 7.7 Hz, H-10), 3.27–3.35
(1H, m, H-3), 4.47–4.49 (1H, m, H-11),
3.82–3.89 (1H, m, H-16), 3.27–3.32 (1H,
m, H-24); for 13C NMR (100 MHz,
CD3OD) spectral data, see Table 1; (2)
FAB-MS: m/z 977 [M 2 H]2, 815
[M 2 162]2, 653 [M 2 162 2 162]2,
491 [M 2 162 2 162 2 162]2; (þ) HR-
ESI-MS: m/z 1001.5290 [M þ Na]þ (cald
for C48H82O20Na, 1001.5297).
3.4 Acid hydrolysis
3.3.1 Curculigosaponin N (1)
Amorphous powder; C42H72O14; ½aꢀ1d5:3 2
Compounds 1 and 2 (2.0 mg) were
dissolved in MeOH (1.0 ml) and 2 M
H2SO4 (1.0 ml) solution and hydrolyzed
under reflux for 2 h, respectively. The
3:41 (c 1.78, MeOH); UV (MeOH) lmax
(log e): 250 (3.16) nm; IR (KBr) nmax
:

Journal of Asian Natural Products Research
411
d

412
A.-X. Zuo et al.
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with 2.0 ml H2O, and extracted with 2.0 ml
EtOAc. The aqueous layer was neutralized
with aqueous Ba(OH)2 and concentrated in
vacuo to give a residue, in which glucose
(from 1 and 2) and rhamnose (from 1) were
identified by comparing with authentic
samples on PC [BuOH–EtOAc–H2O
4:1:5, upper layer, Rf ¼ 0.65 (rhamnose);
BuOH–EtOAc–H2O 4:1:5, upper layer,
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Acknowledgements
This study was financially supported by the
project of Yunnan Science and Technology
Plan (No. 2008IF011), the major projects of
new drugs development of China (No.
2009ZX09102-126), the 973 project of the
Ministry of Sciences and Technology (No.
2009CB941300), and the Open Fund of the
state key laboratory in KIB (No. 0807E31211).
The authors are grateful to the members of the
staff of analytical group of State Key
Laboratory of Phytochemistry and Plant
Resources in West China, Kunming Institute
of Botany, Chinese Academy of Sciences, for
the measurements of all spectra.
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