New triterpenoid saponins from Patrinia scabiosifolia

Article abstract of DOI:10.1080/10286020.2011.653685

Phytochemical investigation of methanol extract from the whole plants of Patrinia scabiosifolia Fisch. resulted in the isolation of three new triterpenoid saponins (1-3) along with twelve known triterpenoids (4-15). The structures of the new compounds wer

Full text of DOI:10.1080/10286020.2011.653685

Journal of Asian Natural Products Research
Vol. 14, No. 4, April 2012, 333–341
New triterpenoid saponins from Patrinia scabiosifolia
Liang Gaoa, Lin Zhangb, Li-Ming Wanga, Jiang-Yun Liua*, Pei-Lie Caia and Shi-Lin Yanga
b
aCollege of Pharmaceutical Science, Soochow University, Suzhou 215123, China; College of
Biomedical Engineering and Instrument Science, Zhejiang University, Hangzhou 310027, China
(Received 15 September 2011; final version received 25 December 2011)
Phytochemical investigation of methanol extract from the whole plants of Patrinia
scabiosifolia Fisch. resulted in the isolation of three new triterpenoid saponins (1–3)
along with twelve known triterpenoids (4–15). The structures of the new compounds
were established as 11a, 12a-epoxy-3-O-b-D-xylopyranosyl-olean-28, 13b-olide (1),
11a, 12a-epoxy-3-O-b-D-xylopyranosyl-(1 ! 3)-a-L-rhamnopyranosyl-(1 ! 2)-b-D-
xylopyranosyl-olean-28, 13b-olide (2), and 3-O-b-D-xylopyranosyl-(1 ! 3)-a-L-
rhamnopyranosyl-(1 ! 2)-b-D-xylopyranosyl oleanolic acid 28-O-b-D-glucopyrano-
side (3) on the basis of various spectroscopic analyses (including different 1D and 2D
NMR spectroscopies and high-resolution electrospray ionization mass spectrometry)
and chemical evidences.
Keywords: Patrinia scabiosifolia; Valerianaceae; triterpenoid saponin
1. Introduction
phytochemical investigations on the
whole plants of Patrinia scabiosifolia
Fisch. collected in China. We have reported
four new saponins along with six known
compounds isolated from the water part of
methanolic extract [3]. Further isolation of
ethanol acetate part of the extract resulted
in three new saponins (1–3, Figure 1),
together with 12 known compounds (4–15)
identified by comparison of their NMR
spectral data with those reported in the
literature [4]. Among them, compounds 1
and 2 possessed an olean-28, 13b-olide
skeleton with 11a, 12a-epoxy, which has
been rarely found in natural sources. The
present study reports the isolation and
structural elucidation of these triter-
penoids, and the cytotoxic activities
of several compounds against HepG-2
cell were also evaluated by 3-[4,5-
dimethylthiazol-2-yl]-2,5-diphenyltetrazo-
lium bromide (MTT) assay in vitro.
The Patrinia genus (Valerianaceae family)
includes about 20 species around the world,
which are widely distributed in Asia and
North America. Of these, more than 10
species grow in Mainland China, many of
which have applications in either tra-
ditional Chinese medicine or folklore
herbs to treat fever, stasis, and inflam-
mation along with detoxication and mobil-
ization of blood circulation [1]. Previous
investigations on the chemical constituents
of Patrinia species have led to the
characterization of several compound
classes including triterpenoids, iridoids,
flavonoids, and sterols. Pentacyclic triter-
penoids are the dominant constituents
within the genus Patrinia and exhibit
oleanane, hederagenin, or ursane skeletons
[2]. As part of our ongoing search for
new natural compounds with interesting
biological activities, we carried out
*Corresponding author. Email: liujiangyun@suda.edu.cn
ISSN 1028-6020 print/ISSN 1477-2213 online
q 2012 Taylor & Francis
http://dx.doi.org/10.1080/10286020.2011.653685
http://www.tandfonline.com

334
L. Gao et al.
29
30
O
20
19
28
21
22
O
O
12
26
18
13
14
11
25
10
17
16
15
Xyl
1
9
6
8
2
27
O
5
3
O
7
HO
HO
4
OR
23
24
1
R = H
2
R = Xyl-(1
3)-Rha-
O
Xyl
O
O
O
HO
HO
O
HO
O
HO
HO
H3C
HO
Rha
O
HO
Glc
O
O
HO
HO
HO
OH
Xyl'
Figure 1. Chemical structures of compounds 1–3.
2. Results and discussion
13C NMR spectrum (seven sp3 carbons at
dC: 16.5, 17.4, 19.0, 20.5, 23.6, 28.0, and
33.2) (Table 1). The carbonyl carbon signal
at dC 179.0 together with the quaternary
carbon at dC 87.7 indicated that the aglycon
possessed a 28, 13b-lactone [5]. Two
oxygen-bearing methylenic protons at dH
3.17 (1H, d, J ¼ 4.0 Hz) and 3.32 (1H,
d, J ¼ 4.0 Hz), and two corresponding
oxygen-bearing methine carbons at dC
52.9 and 57.4 indicated the presence of a
ternary oxygen ring. After extensive 2D
NMR analysis, the aglycon of 1 was
Compound 1 was obtained as a white
amorphous powder. The positive HR-ESI-
MS exhibited a pseudomolecular ion peak
at m/z 625.3716 [M þ Na]þ, correspond-
ing to the molecular formula of C35H54O8.
The IR spectrum of 1 exhibited absorptions
at 3423 cm21 (ZOH), 1769 cm21 (ester
carbonyl), and 871 cm21 (epoxy). The H
1
NMR spectrum of 1 showed signals for
seven tertiary methyl groups at dH 1.37,
1.32, 1.22, 1.06, 1.01, 0.98, and 0.88 (each
3H, s), coupled with information from the

Journal of Asian Natural Products Research
335

336
L. Gao et al.
characterized as 11a, 12a-epoxy-olean-28,
13b-olide, referring to the reported data of
11a, 12a-epoxy-3-O-b-D-glucuronopyra-
nosyl-olean-28, 13b-olide [6]. In HMBC
spectrum of 1 (Figure 2), the correlations of
11, 12-epoxy ring could be observed from
H-11 (dH 3.17) to C-10 (dC 36.6) and C-12
(dC 57.4), from H-12 (dH 3.32) to C-11
(dC 52.9) and C-14 (dC 41.0), from H-9
(dH 1.79) to C-11 (dC 52.9), and from H-18
(dH 2.61) with C-12 (dC 57.4); the
correlations of 28, 13-lactone ring could
be observed from H-15 (dH 1.09), H-18
(dH 2.61) and H-27 (dH 1.32) to C-13
(dC 87.7), from H-18 (dH 2.61) to C-28
(dC 179.0), respectively, thus confirming
the moiety connection of the aglycon.
Further, NOE relationships (Figure 2)
between H-11 at dH 3.17 and Me-25 at dH
1.01/Me-26 at dH 1.22 (Me-26), and
between H-12 at dH 3.32 and Me-26 at dH
1.22 convinced the a-configuration of the
epoxy ring. The presence of one sugar
could be deduced from its anomeric proton
signal at dH 4.90 (1H, d, J ¼ 7.5 Hz) and
anomeric carbon signal at dC 107.8
(Table 2). The b-anomeric configuration
of D-xylose unit was determined from its
3JH1,H2 coupling constants(7.5 Hz) [7]. The
linkage position of b-D-xylose was shown
to be at C-3 of the aglycon by detecting a
correlation from the anomeric proton at dH
4.90 (Xyl-1) to C-3 at dC 88.5 in the HMBC
spectrum (Figure 2). On acid hydrolysis
with 2 M CF3COOH, 1 afforded sugar
moiety that was identified as D-xylose
based on the gas chromatography (GC)
analysis of its chiral derivative [8]. The
structure of 1 was finally established as
11a, 12a-epoxy-3-O-b-D-xylopyranosyl-
olean-28, 13b-olide.
871 cm21 (epoxy). A detailed comparison
of the 1H and 13C NMR chemical shifts of 2
with those of 1 revealed the same aglycon
moiety for both compounds, with differ-
ences of sugar moieties (Tables 1 and 2).
On acid hydrolysis with 2 M CF3COOH, 2
afforded sugar moieties as L-rhamnose and
D-xylose in the ratio of 1:2 based on the GC
analysis of its chiral derivative [8]. The 1H
and 13C NMR spectra of 2 exhibited three
sugar anomeric protons at dH 6.67 (1H, br
s), 5.46 (1H, d, J ¼ 7.5 Hz), 4.91 (1H, d,
J ¼ 7.0 Hz), with three anomeric carbon
signals at dC 101.7, 107.8, and 106.3
correspondingly. The methyl signal at dH
1.74 (3H, d, J ¼ 6.0 Hz) and dC 18.7 also
indicated the presence of a rhamnose. The
glycosylation shift of C-3 (dC 88.4)
revealed that 2 was a 3-monodesmosidic
glycoside. The identities of the oligo-
saccharide sequence were determined by
a combination of HSQC, HMBC, and
NOESY experiments (Table 2). Following
HMBC correlations could be observed:
from H-1 of the xylose (dH 4.91, inner) to
C-3 (dC 88.4) of the aglycon, H-1 (dH 6.67)
of rhamnose to C-2 (dC 77.2) of the xylose
(inner), and H-1 of another xylose (dH 5.46)
(terminal) to C-3 (dC 83.3) of rhamnose
(Figure 2). The b-configuration at the
anomeric positions of the xylose units was
3
determined from its JH1,H2 coupling
constants (7.0–8.0 Hz) [7], and the chemi-
cal shift of C-5 (dC 69.8) indicated the usual
a-configuration for the rhamnose unit [9].
These configurations were also confirmed
from the NOE relationships between H-1
and H-3 and between H-1 and H-5 of
monosaccharide moieties (Figure 2) [10].
The structure of 2 was thus established as
11a, 12a-epoxy-3-O-b-D-xylopyranosyl-
(1 ! 3)-a-L-rhamnopyranosyl-(1 ! 2)-b-
D-xylopyranosyl-olean-28, 13b-olide.
Compound 3 was obtained as a
white amorphous powder. The positive
HR-ESI-MS exhibited a pseudomolecular
ion peak at m/z 1051.5448 [M þ Na]þ,
corresponding to the molecular formula
of C52H84O20. The IR spectrum of 3
Compound 2 was obtained as a white
amorphous powder. The positive FTICR-
HR-ESI-MS exhibited a pseudomolecular
ion peak at m/z 903.4716 [M þ Na]þ,
corresponding to the molecular formula
of C46H72O16. The IR spectrum of 2
exhibited absorptions at 3418 cm21
(ZOH), 1770 cm21 (ester carbonyl), and

Journal of Asian Natural Products Research
337
29
30
O
20
19
28
H
O
21
22
O
H
25
12
26
H
1
18
H
15
17
16
11
13
14
9
Xyl
10
2
8
5
27
O
O
3
7
HO
HO
4
6
OH
23
24
H
NOESY
HMBC
29
30
O
20
19
21
22
H
O
O
28
H
25
12
H
18
13
14
17
16
26
11
H
15
1
4
9
10
2
3
8
Xyl
O
27
O
O
5
O
7
6
HO
HO
23
Rha
HO
24
H3C
HO
H
H
O
NOESY
HMBC
O
HO
HO
OH
Xyl
H
12
13
O
Xyl
28
3
O
O
O
HO
HO
O
HO
O
HO
HO
H3C
HO
Rha
O
H
H
HO
Glc
O
O
HO
HO
H
HO
OH
H
Xyl′
HMBC
Figure 2. Key HMBC and NOESY correlations for compounds 1–3.

338
L. Gao et al.
Table 2. The 1H and 13C NMR spectral data for sugar moieties of compounds 1–3 in pyridine-d5
(J in Hz).
1
2
3
No.
13C
1H
13C
1H
13C
1H
3-Xyl
1
2
3
4
5
107.8
75.6
78.8
71.4
67.2
4.90 d (7.5)
4.11 br d (7.5)
4.24 m
4.32 m
4.46 m
3.85 d (10.5)
106.3
77.2
79.9
71.6
67.2
4.91 d (7.0)
4.36 m
4.28 m
4.26 m
4.41 m
3.78 d (10.0)
106.3
77.0
80.0
71.7
67.2
4.91 d (7.5)
4.35 m
4.27 m
4.25 m
4.40 m
3.78 d (10.5)
Rha
1
2
3
4
101.7
72.1
83.3
73.2
69.8
18.7
6.67 br s
5.09 br s
4.89 m
4.62 m
4.87 m
1.74 d (6.0)
101.6
72.1
83.2
73.2
69.8
18.7
6.67 br s
5.08 br s
4.89 m
4.61 br d (9.5)
4.86 dd (6.0, 9.5)
1.72 d (6.0)
5
6
Xyl0
1
2
3
4
5
107.8
75.8
78.7
71.3
67.6
5.46 d (7.5)
4.17 br d (7.5)
4.30 m
4.34 m
4.43 br d (9.0)
3.82 d (9.0)
107.8
75.8
78.7
71.3
67.7
5.46 d (7.5)
4.16 br d (7.5)
4.28 m
4.31 m
4.42 m
3.82 d (11.0)
28-Glc
1
2
3
4
5
6
95.9
74.3
79.1
71.3
79.5
62.4
6.42 d (8.0)
4.28 m
4.35 m
4.43 m
4.12 m
4.54 m, 4.49 m
exhibited absorptions at 3416 cm21
(ZOH), 1750 cm21 (ester carbonyl), and
1641 cm21 (double bond). The NMR
spectra of compound 3 showed character-
istic signals of a triterpenoid saponin.
Seven tertiary methyl groups at dH 0.96,
0.97, 0.99, 1.17, 1.32, 1.35, and 1.47, and
corresponding seven sp3 carbons at dC 15.8,
23.8, 33.3, 17.6, 17.5, 26.3, and 28.3 could
be observed, together with one trisubsti-
tuted olefinic proton at dH 5.50 (1H, br s)
and two sp2 olefinic carbons at dC 123.0 and
144.3, which indicated the typical olean-
12-ene aglycon signals (Table 1). The
chemical shifts of C-3 (dC 88.7) and C-28
(dC 176.6) revealed that 3 was a bisdesmo-
sidic glycoside. The 1H and 13C NMR
spectra of 3 exhibited four sugar anomeric
protons at dH 6.67 (br s), 6.42 (d,
J ¼ 8.0 Hz), 5.46 (d, J ¼ 7.5 Hz), and
4.91 (d, J ¼ 7.5 Hz), and carbons at dC
95.9, 101.6, 106.3, and 107.8, which
ascribed to remaining 22 oligosaccharide
carbon signals of 3 (Table 2). The methyl
carbon signal at dC 18.7 and the doublet
methyl proton signal at dH 1.72 (3H, d,
J ¼ 6.0 Hz) indicated the presence of a
6-deoxy sugar. The identities of the
oligosaccharide sequence were determined
by a combination of COSY, HSQC,
HMBC, and NOESY experiments. The
oligosaccharide connected to C-3 of the
aglycon was elucidated as C-3-Xyl-
(1 ! 3)-Rha-(1 ! 2)-Xyl, same as that of

Journal of Asian Natural Products Research
2. A disaccharide part at C-28 of the 3. Experimental
339
aglycon was established by the HMBC
correlation from the H-1 (dH 6.42) of
glucose to C-28 (dC 176.6) of the aglycon
(Figure 2). On acid hydrolysis with 2 M
CF3COOH, 3 afforded sugar moieties that
were identified as L-rhamnose, D-xylose,
and D-glucose in the ratio of 1:2:1 based on
the GC analysis of its chiral derivative [8].
On the basis of the above results, the
structure of 3 was finally established as 3-
O-b-D-xylopyranosyl-(1 ! 3)-a-L-rham-
nopyranosyl-(1 ! 2)-b-D-xylopyranosyl
oleanolic acid 28-O-b-D-glucopyranoside.
Moreover, 12 known triterpenoid
compounds were also isolated and ident-
ified by comparing their NMR and MS
spectral data with those reported in the
literature. They are assigned to be oleanoic
acid (4) [3], 2a-hydroxyoleanoic acid (5)
[3], 3a-ursoloic acid (6) [4], 3-hydroxy-
olean-11-oxo-12-en-28-oic acid (7) [4],
3, 11-dioxo-olean-12-en-28-oic acid (8)
[4], 29-hydroxy-3-oxo-olean-12-en-28-oic
acid (9) [4], 3b, 12a-dihydroxy-oleanan-
13b, 28-olide (10) [4], oleanolic acid 28-
O-b-D-glucopyranosyl-(1 ! 6)-b-D-glu-
copyranosyl ester (11) [4], 3-O-b-D-
xylopyranosyl oleanolic acid 28-O-b-D-
glucopyranosyl ester (12) [4], 3-O-a-L-
rhamnopyranosyl-(1 ! 2)-b-D-xylopyra-
nosyl-oleanolic acid 28-O-b-D-glucopyr-
anosyl ester (13) [4], oleanolic acid 3-O-b-
D-xylopyranoside (14) [4], and oleanolic
acid 3-O-a-L-rhamnopyranosyl-(1 ! 2)-
b-D-xylopyranoside (15) [4]. Among
them, seven compounds 6–10, 12, and
13 were isolated from this genus for the
first time.
3.1 General experimental procedures
Optical rotations were measured on a
Perkin-Elmer PL341 polarimeter, and IR
spectra were recorded on a Perkin-Elmer
938G instrument (Perkin-Elmer, Inc.,
Waltham, Massachusetts, USA). The
NMR spectra were recorded on a Varian
500 MHz NMR System (Varian, Inc.,
Palo Alto, California, USA). HR-ESI
mass spectra were recorded with Finnigan
LCQ-Deca (for compounds 1–3; Thermo-
Finnigan, Inc., San Jose, California, USA)
and Varian 7.0 T FTICR (for compound 2;
Varian, Inc., Palo Alto, California, USA)
mass spectrometers. GC analysis was
carried out on a Shimadzu GC-14C gas
chromatograph (Shimadzu Corporation,
Kyoto, Japan) using a Rtx-1 capillary
column (30 m £ 0.25 mm, i.d.); flame
ionization detector; detector temperature,
2508C; injection temperature, 2508C;
initial temperature was at 1008C then raised
to 1808C at the rate of 108C/min; and from
1808C to 2308C at the rate of 38C/min;
carrier gas, N2 (0.8 ml/min). HPLC separ-
ations were performed on a Shimadzu LC-
20A series apparatus with an SPD-20A UV
detector
(Shimadzu
Corporation),
equipped with a 250 £ 20 mm i.d. prepara-
tive Cosmosil AR-II C18 column (Nacalai
Tesque, Inc., Kyoto, Japan); Medium
pressure liquid chromatography was per-
formed on a Lisure HP Purifier apparatus
with a UV detector (Lisure science
corporation, Suzhou, China). Silica gel
(200–300 mesh); (Qingdao Haiyang
Chemical, Inc., Qingdao, China) and
Cosmosil C18 reversed-phase silica gel
(75 mm, Nacalai tesque corporation)
were used for column chromatography.
The spray reagent for saponins was
ethanolic H2SO4 (10%).
Selected compounds 2, 3, and 8–11
were evaluated for their cytotoxic activi-
ties against human hepatoma cell lines
(HepG-2) by the MTT assay in vitro, while
compound 1 was not tested due to limited
amount. Compound 2 exhibited moderate
cytotoxicity to HepG-2 cells with IC50
value of 28.5 mM, while other compounds
did not show cytotoxicity at a high sample
concentration of 100 mM.
3.2 Plant material
The whole plants of P. scabiosifolia were
collected from Guangnan County, Yunnan

340
L. Gao et al.
3.3.1 Compound 1
Province, China, in August 2008, and the
botanical origin of the material was
identified by Prof. Lin Zhang, College of
Biomedical Engineering & Instrument
Science, Zhejiang University, Hangzhou,
China. The voucher specimens (No.
PS080726) are deposited at the Depart-
ment of Natural Medicinal Chemistry,
College of Pharmaceutical Science, Soo-
chow University, Suzhou, China.
20
White amorphous powder; ½aꢀD 22.1
(c ¼ 0.22, MeOH); IR (KBr) nmax 3423
(ZOH), 2936, 2871 (CH), 1769 (CvO),
1
871 (epoxy) cm21; H NMR (pyridine-d5,
500 MHz) and 13C NMR (pyridine-d5,
125 MHz) spectral data: see Tables 1 and
2; HR-ESI-MS: m/z 625.3716 [M þ Na]þ
(calcd for C35H54O8Na, 625.3711).
3.3.2 Compound 2
3.3 Extraction and isolation
20
White amorphous powder; ½aꢀD 28.2
The air-dried plants (2.5 kg) were
extracted with 95% aqueous methanol
(v/v) for three times (8 liters, 1.5 h each) at
room temperature. After evaporation, the
residue was suspended in H2O and
partitioned using EtOAc (6 £ 4 liters).
The EtOAc fraction (81.3 g) was subjected
to vacuum liquid chromatography on silica
gel (30 £ 25 cm) using a stepwise gradient
elution of CHCl3–MeOH (100:1 – 70:30)
to afford 11 subfractions (PE-1–11). PE-6
(5.32 g) was subjected to silica gel column
with gradient elution of CHCl3–MeOH
(30:1–25:1), followed by medium press-
ure liquid chromatography over ODS-C18
column eluted with MeOH-H2O (75:25–
95:5), and further purified by Sephadex
LH-20 with CHCl3–MeOH (1:1) to yield 1
(10.2 mg). PE-10 (5.46 g) was further
chromatographed on silica gel column,
followed by preparative HPLC (MeOH–
H2O 78:12; flow rate 2 ml/min; UV
210 nm detection) to yield 13 (29.7 mg),
2 (18.7 mg), and 11 (17.5 mg), respect-
ively. PE-11 (1.02 g) was further chro-
matographed on silica gel column eluted
with CHCl3–MeOH (9:1 – 8:2) to yield 3
(21.4 mg). By similar separation pro-
cedures, compound 8 (22.3 mg) from PE-
1 (12.17 g), 5 (16.7 mg), 6 (13.1 mg), 9
(18.4 mg) and 10 (16.6 mg) from PE-2
(3.15 g), 4 (240 mg) and 7 (12.5 mg) from
PE-3 (4.23 g), 14 (24.7 mg) from PE-7
(11.49 g), 12 (33.6 mg) and 15 (20.3 mg)
from PE-9 (5.03 g) were also obtained,
respectively.
(c ¼ 0.19, MeOH); IR (KBr) nmax 3418
(ZOH), 2933, 2866 (CH), 1770 (CvO),
871 (epoxy) cm21; H NMR (pyridine-d5,
500 MHz) and 13C NMR (pyridine-d5,
125 MHz) spectral data: see Tables 1 and
2; HR-ESI-MS: m/z 903.4593 [M þ Na]þ,
FTICR-HR-ESI-MS: m/z 903.4716
[M þ Na]þ (calcd for C46H72O16Na,
903.4713).
1
3.3.3 Compound 3
20
White amorphous powder; ½aꢀD 27.7
(c ¼ 0.20, MeOH); IR (KBr) nmax 3416,
1
2943, 1750, 1641, 1073 cm21; H NMR
(pyridine-d5, 500 MHz) and 13C NMR
(pyridine-d5, 125 MHz) spectral data: see
Tables
1 and 2; HR-ESI-MS: m/z
1051.5448 [M þ Na]þ (calcd for
C52H84O20Na, 1051.5488).
3.3.4 Acid hydrolysis and GC analysis of
1–3
Each compound (3 mg) was dissolved
in 2 M CF3COOH (2 ml) and heated at
1208C for 4 h in a sealed tube. The reaction
mixture was extracted with EtOAc
(2 ml £ 3). Each remaining aqueous layer
was concentrated to dryness to give a
residue. The residue was dissolved in
pyridine (1 ml), and then L-cysteine methyl
ester hydrochloride (2 mg) was added to the
solution. The mixture was heated at 608C
for 1 h, and trimethylchlorosilane (0.5 ml)
was added, followed by heating at 608C for

Journal of Asian Natural Products Research
341
tute of Organic Chemistry, Chinese Academy
of Science for the NMR measurements. This
work was supported by a project funded by the
Priority Academic Program Development of
Jiangsu Higher Education Institutions (PAPD).
30 min. Then, the solution was concen-
trated to dryness and dissolved in water
(1 ml £ 3), followed by extraction with
n-hexane (1 ml £ 3). The hexane extract
was subjected to GC analysis [8]. The
absolute configurations of the monosac-
charides were confirmed to be L-rhamnose,
D-xylose, and D-glucose by comparison
of the retention times of monosaccharide
derivatives with those of standard
samples: L-rhamnose (8.32 min), D-xylose
(10.55 min), and D-glucose (15.23 min),
respectively.
References
[1] Editorial Committee of Nanjing Univer-
sity of Traditional Chinese Medicine, The
Dictionary of Chinese Herbal Medicine
(Shanghai Science and Technology Pub-
lishing House, Shanghai, 2006), p. 1914.
[2] Y.F. Li, F.C.H. Lou, Y.P. Tang, and J.H.
Wang, Nat. Prod. Res. Dev. 13, 71 (2001).
[3] L. Gao, L. Zhang, N. Li, J. Liu, P. Cai, and
S. Yang, Carbohyd. Res. 346, 2881 (2011).
[4] L. Gao, L. Zhang, J. Liu, P. Cai, and S.
Yang, Chin. Tradit. Herb. Drugs 42, 1477
(2011).
[5] L.W. Fu, S.J. Zhang, J. Sakai, T.
Hasegawa, T. Mitsui, S. Oka, K. Hirose,
and M. Ando, J. Nat. Prod. 68, 198 (2005).
[6] M. Kuroda, T. Aoshima, M. Haraguchi,
M.C. Marx Young, H. Sakagami, and Y.
Mimaki, J. Nat. Prod. 69, 1606 (2006).
[7] O. Safir, S. Fkih-Tetouani, N. De Tom-
masi, and R. Aquino, J. Nat. Prod. 61, 130
(1998).
3.4 MTT cytotoxicity assay
The bioassay was carried out according to
the method described by Chen et al. [11].
Compounds 2, 3, and 8–11 were evaluated
against the HepG-2 human hepatoma cell
lines, and compound 2 was cytotoxic to
HepG-2 cells with IC50 value of 28.5 mM.
Supporting information
Supporting information of compounds
1–3 can be found in the online version
of this article.
[8] Q. Chen, J.G. Luo, and L.Y. Kong, Chem.
Pharm. Bull. 58, 412 (2010).
[9] R. Kasai, M. Okihara, J. Asakawa, K.
Mizutani, and O. Tanaka, Tetrahedron 35,
1427 (1979).
[10] Q. Zheng, K. Koike, L.K. Han, H. Okuda,
and T. Nikaido, J. Nat. Prod. 67, 604
(2004).
[11] C.R. Chen, Y.W. Liao, H.T. Wu, W.L.
Shi, C.Y. Tzeng, S.Z. Yang, C.E.
Hernandez, and C.I. Chang, Chem.
Pharm. Bull. 58, 408 (2010).
Acknowledgements
We are grateful to Prof. Xue-Dong Yang of the
Department of Pharmaceutical Science and
Technology, Tianjin University, for the HR-
ESI-MS measurement and Mrs Li-Ping Shi of
the Instrumentation Center of Shanghai Insti-

Copyright of Journal of Asian Natural Products Research is the property of Taylor & Francis Ltd and its content
may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express
written permission. However, users may print, download, or email articles for individual use.