532
J.-G. Luo et al.
3.3 Extraction and isolation
The roots of G. pacifica (8.9 kg) were
ground into powders and then extracted
with 70% aqueous EtOH (v/v) three times
(10 l, 2 h each) under reflux. After
evaporation, the residue was suspended
in water and partitioned by EtOAc,
n-BuOH, and water. The EtOAc portion
(60 g) was fractionated by silica gel
column (100–200 mesh), which was
eluted with CHCl3/MeOH with gradually
increasing polarity to give five fractions
(fractions 1–5); fraction 3 (500 mg,
CHCl3/MeOH, 10:1, v/v) was further
subjected to repeated silica gel column
with CHCl3/MeOH gradually to yield pure
2 (105 mg), 3 (7 mg), and 4 (4 mg),
respectively. The n-BuOH-soluble portion
(268 g) was fractionated by MCI gel,
which was eluted with MeOH/H2O
(0:100, 30:70, 50:50, 70:30, and 100:0,
v/v) to give five fractions (fractions 1–5);
fraction 5 (180 mg, MeOH/H2O, 100:0,
v/v) was further purified by repeated silica
gel column (CHCl3/MeOH, 3:1, v/v) to
afford pure 1 (28 mg).
Figure 2. Key HMBC correlations for
compound 1.
3. Experimental
3.1 General experimental procedures
Optical rotations were measured with a
JASCO P-1020 polarimeter. IR (KBr
disks) spectra were recorded by Bruker
Tensor 27 spectrometer. 1D and 2D NMR
spectra were recorded at ACF-500 NMR
instrument (1H: 500 MHz, 13C: 125 MHz),
with TMS as an internal standard. Mass
spectra were obtained on an MS Agilent
1100 Series LC/MSD Trap mass spec-
trometer (ESI-MS) and a Micro Q-TOF
MS (HR-ESI-MS), respectively. TLC was
performed on a precoated silica gel G
(Qingdao Haiyang Chemical Co. Ltd,
Qingdao, China), and detection was
achieved by 15% H2SO4 –EtOH for
triterpenoids. Silica gel H (Qingdao
Haiyang Chemical Co. Ltd) and Sephadex
LH-20 (Pharmacia, Amersham Bios-
ciences, Uppsala, Sweden) were used for
column chromatography.
3.3.1 3b-O-Sulfate gypsogenin 28-O-b-
D-glucopyranosyl ester (1)
A white amorphous powder (MeOH);
[a]2D5 þ 5.95 (c 0.01; MeOH); IR (KBr)
n
max: 3418, 2948, 1735, 1639, 1229,
1073 cm21. H NMR (500 MHz, C5D5N)
and 13C NMR (125 MHz, C5D5N) spectral
data, see Table 1. ESI-MS m/z: 711
[M 2 H]2, 730 [M þ NH4]þ, 735
[M þ Na]þ; HR-ESI-MS m/z: 711.3433
[M 2 H]2 (calcd for C36H55O12S,
711.3420).
1
3.2 Plant material
The roots of G. pacifica were collected
from Xifeng region, Liaoning Province,
China, in October 2005. The botanical
origin of material was identified by Prof.
Minjian Qin, Research Department of
Traditional Chinese Medicinal Resources,
China Pharmaceutical University, and the
voucher specimens (No. 051020) have
been deposited at the Department of
Natural Medicinal Chemistry, China Phar-
maceutical University, Nanjing, China.
3.3.2 3b-O-Sulfate gypsogenin (2)
A white amorphous powder (MeOH);
[a]2D5 þ 42.03 (c 0.08; MeOH); IR (KBr)
n
max: 3442, 2948, 1690, 1638, 1244,
1078 cm21. H NMR (500 MHz, CD3OD)
and 13C NMR (125 MHz, CD3OD) spec-
tral data, see Table 1. ESI-MS m/z: 549
1