全 文 ：Polyphenols from Vitis thunbergii Sieb.et Zucc.
DOU De-qiang1＊ , REN Jie1 , Maggie Cooper3 , HE Yue-hua2 , PEI Yu-ping1 , Yoshiaki Takaya2 ,
Masatake Niwa
2 , CHEN Ying-jie1 , YAO Xin-sheng1 , and ZHOU Ren-ping3
(1 .Department of Natural Products Chemistry , Shenyang Pharmaceutical University , Shenyang 110016 , China;
2 .Faculty of Pharmacy , Meijo University , Japan;3 .Department of Chemical Biology , Rutgers University , USA)
Abstract:Aim　Isolation and structural elucidation of the constituents from the aerial part of Vitis thunbergii.Meth-
ods　To isolate chemical constituents , column chromatography and HPLC were used.Physico-chemical characterization and
spectroscopic analysis were employed for structural identification.Results　Eleven polyphenols were isolated and identified.
Conclusion　Compound 1 is a new compound and its structure was characterized to be 3 , 5-dimethoxyl-4-hydroxyl-phenyl-
propanol-9-O-β-D-gly copyranoside.
Key words:Vitis thunbergii;phenylpropanol glycoside;polyphenol
CLC number:R284.2　　　Document code:A　　　Article ID:1003-1057(2003)02-057-03
　　Vitis thunbergii Sieb.et Zuce.is mainly distributed
in China , Korea , and Japan.In Chinese folklore both the
aerial part and roots of this plant are used as drugs in the
treatment of dysentery , rheumatism , and tuberculosis.
Research indicated that the fruit contains 10%sugar , or-
ganic acid , and vitamins[ 1] .Recent research showed that
Vitis vinifera , which shares the same genus with the title
plant , contains a high content of polyphenols possessing
potential cancer-chemopreventive and antioxidant
activities
[ 2] .In order to explore the active constituents
and provide chemotaxonomic information , the chemical
constituents of Vitis thunbergii have been systematically
studied.From its aerial part 45 compounds were isolated.
Out of them 11 compounds were identified;compound 1
is a new compound.This paper deals with the isolation
and structural elucidation of the new compound.
Experimental
Materials and methods
[α] D was recorded at 589 nm on a JASCO P-1020
spectrometer(cell length 100 mm).CD spectroscopy was
conducted on JASCO J-600.1H NMR and 13C NMR data
were obtained on JEOL J-500(500 MHz), and chemical
shifts were given inδwith their solvents(CD3OD or Ace-
Received date:December 29, 2002
＊Corresponding author:Tel 024-23882129
E-mail doudeqiang@yahcc .com
tone-d6)as an internal standard.FAB-MS was taken on
HX-110 mass spectrometer , and ESI-MS on LCQ ESI-MS
spectroscopy.
The aerial part of Vitis thunbergii was collected in
Toyoake , Aishi prefecture , Japan , in September 2000.
The specimens were stored in the Faculty of Pharmacy ,
Meijo University.
Column chromatography was performed on Amberlite
XAD-2(Olugana Corporation , Tokyo , Japan), Silica gel
BW-820 MH (Fuji Division), and Sephadex LH-20
(Amersham Pharmacia Biotech , Sweden).TLC was con-
ducted on Silica gel 60 F254 with chloroform-MeOH in dif-
ferent ratios as development solvents and Rp-18 F254(Mer-
ck)with mixture of MeOH-H2O;the spot was detected by
spraying the TLC with dilutedH2SO4 followed by heating.
MPCC over Rp-18(Develosil ODS , Nomura Chemi-
cal Co.Ltd.;Size:25 mm×350 mm)was run by a #
GK12 mini pump (Oriental Motor Corporation , Japan),
and was eluted with MeOH-H2O in gradient.
Preparative HPLC was carried out on a LC-10(Ja-
pan Analytical Industry Co.Ltd.)equipped with RI and
UV detectors , using Develosil C8-5 and C-30-UG-5
(C30 , Nomura Chemical Co.Ltd., Size 20 mm×250
mm)as well as C18 column(waters).
Extraction and isolation
The fresh aerial part of Vitis thunbergii (18 kg)was
extracted with MeOH 3 times to afford 953 g of MeOH ex-
57Journal of Chinese Pharmaceutical Sciences 2003 , 12(2)
tract.This was suspended in 10%MeOH and partitioned
with EtOAc and n-BuOH , respectively , to yield EtOAc
extract(299 g)and BuOH extract(257 g).
A portion of n-BuOH extract.(140 g)was suspend-
ed in water and subjected to Amberlite XAD-2 column
(2 000mL.Elution with water , 30%MeOH , MeOH and
acetone followed by evaporation in vacuum provided the
corresponding eluates:H2O eluate (75 g), 30%MeOH
eluate (5.8 g), MeOH eluate(49 g), and acetone eluate
(6 g).
The MeOH eluate was subjected to column chroma-
tography over silica gel(820 H , 500 g)and eluted with
CHCl3-MeOH in gradient to afford Fr.1 (0.6 g ,
CHCl3), Fr.2(1 g , 10%MeOH ), Fr.3 (2 g , 20%
MeOH), Fr.4(12 g , 20%MeOH), Fr.5(9 g , 30 %
MeOH), Fr.6 (5 g , 40%MeOH), Fr.7 (6 g , 50%
MeOH), and Fr.8(12 g , MeOH).
MPCC of Fr.4 (12 g)over silica gel (200 g ,
45 mm ×450 mm)using a gradient solvent system of
CHCl3-MeOH (10∶1 to 2∶1)gave 10 fractions(Fr.41 to
Fr.410).Further chromatography of Fr.44 (1.6 g ,
CHCl3-MeOH =8∶1)over Develosil ODS , eluted with
MeOH-H2O in gradient (20% MeOH , 40% MeOH ,
60% MeOH and MeOH), gave 9 additional fractions
(Fr.441 to Fr.449).A reversed phase HPLC (C18 col-
umn)of Fr.442 using MeOH-H2O (25∶75), 2 mL·
min
-1;and RI detector , furnished 2 (5 mg), 3 (10
mg), 4(7mg), 5(16mg)and 1(10 mg)in accordance
with the detection sequence of peaks;Fr.447 was chro-
matographed by HPLC(C8-5;MeOH-H2O=43∶57 , 3.5
mL·min-1;UV 220 nm detection)to afford 6(8.8 mg),
7(2.0 mg), 8(3.2 mg)and 9(16 mg).
Fr.5 (9 g)was subjected to column chromatography
over silica gel eluted with CHCl3-MeOH-H2O (69∶27∶4)to
give 6 fractions(Fr.51 to Fr.56).A reversed phase MPLC
(Develosil ODS)of Fr.53 , eluted withMeOH-H2O in gradi-
ent(20%MeOH to 100%MeOH)furnished 12 subfractions
(Fr.531 to Fr.5 312).Fr.534(30%MeOH eluate)was
further purified by HPLC (C-30-UG-5 column , MeOH-H2O
(4∶6), 3mL·min-1;UV 220nm)to give compound 10(4.2
mg).HPLC of Fr.539(50%MeOH eluate;C8-5 column;
MeOH-H2O(5∶5), 3mL·min-1;220 nm detection)afford-
ed compound 11(5.3 mg).
Identification
Compound 1 　White powder;[ α] D:-24.8(c
0.3;24℃);UV(nm , logε):208.4 (4.51), 276.8
(3.32);IR(cm-1):3 421 , 1 619 , 1 517 , 1 457;HR-
FAB-MS(positive , m z):Found:397.1573 , Calcd for
C17H26O9[M] +:397.1577.ESI-MS (m z):397 [M+
Na] + , 392 [ M+H2O] +;1H NMR and 13C NMR are
listed in Table 1.
Table 1　1H NMR and 13 C NMR data of 1 in CD3OD
Carbon δc δH
1 134.1 s
2 , 6 107.0 d 6.50(2H , s)
3 , 5 149.2 s
4 134.7 s
7 32.9 t 2.64(2H , t , J=7.2 Hz)
8 33.1 t 1.90(2H , m)
9 69.9 t 3.91(1H , m , Ha);3.53(1H , m , Hb)
Glc
1′ 104.5 d 4.26(1H , d , J=7.8 Hz)
2′ 75.2 d 3.20(1H , dd , J=7.8 , 8.0 Hz)
3′ 78.2 d 3.35(1H , dd, J=8.0 , 8.7 Hz)
4′ 71.7 d 3.30(hidden)
5′ 77.8 d 3.25(1H , m)
6′ 62.8 t 3.85(1H , dd , J=2.3 , 11.8Hz);3.67(1H ,
dd , J=5.5 , 11.8 Hz)
-OCH3 56.8 q 3.82(6 H , s)
Figure 1　Correlations in 1H-1H COSY of 1
　　By comparison of their [α] D , 1H NMR and 13C NMR
and MS spectra with the data in the references , com-
pounds 2-11were identified as syringin(2), dihydrosy-
ringin (3), catechin (4)[ 5] , O-hydroxybenzylglycoside
(5), miyabenol C (6), cis miyabenol C (7)[ 6] , α-
viniferin(8), (+)-ε-viniferin(9), resveratrol-3-O-β-D-
glucopyranoside (10)[ 7] and ampelopsin C(11)[ 8] .
Results and Discussion
Compound 1 was obtained as an amorphous powder
with a strong OH absorption in its IR spectrum , which
gave D-glucose on acid hydrolysis.The molecular formula
C17H26 O9 was drawn from HRFAB-MS , ESI-MS and
13
C NMR spectra.The ESI-MS showed quasi-molecular
58 Journal of Chinese Pharmaceutical Sciences 2003 , 12(2)
ion peak at m z 397 [M+Na] + and 392 [M+H2O] +.
The
13
C NMR spectra showed 17 carbon signals;the
DEPT technique suggested that the seventeen
13
C signals
were comprised of 2 methyls , 4 methenes , 7 methines ,
and 4 quarternary carbon signals.Among them a set of
signals due to a β-D-glucopyranosyl unit at δ104.5 ,
75.2 ,78.2 ,71.7 ,77.8 ,62.8 was also observed , disclos-
ing 1 as a mono-β-D-glucopyranoside.This was confirmed
by the inspection of anomeric signal δ4.26 in the 1H
NMR spectrum.The coupling constant(J=7.8 Hz)of
the anomeric H indicated the β-configuration of the glu-
cosyl group.
The structure of aglycone can be elucidated as fol-
lows.The fact that the 1H NMR signal atδ6.50(2H , s)
correlated with carbon signal at δ107.0 in the HMQC
spectrum of 1 coupled with signals atδ134.1 , 134.7 and
149.2 (two magnitude equivalent carbon)revealed the
presence of a 1 , 3 , 4 , 5-tetrasubstituted phenyl group in
the aglycone.In the 1H NMR spectrum the characteristic
1
H NMR signals of two methoxyls observed at δ3.82
(6H , s)indicated that the two methoxyls were at the
symmetrical C-3 and C-5 of the phenyl group.According
to its molecular formula , a hydroxyl group must exist in
the aglycone and be located at C-4.The propanol group
was suggested to be linked to C-1 by the analysis of HM-
BC spectra of 1.Long-range correlations were observed
between H-7 and C-2 , 6 , 8 , 9 , between H-2 , 6 , 9 , and
C-7 in the HMBC spectra.Therefore , the structure of ag-
lycone was elucidated to be 3 ,5-dimethoxyl-4-hydroxyl-1-
phenyl propanol.
By glycosidation shift rule analysis , the sugar should
be linked to C-9 of aglycone in compound 1 as compared
with the
13
C NMR data of propanol group.Moreover ,
long-range correlation was observed between anomeric H
and C-9 of aglycone in the HMBC spectrum.Following
these analyses , 1 was elucidated to be 3 , 5-dimethoxyl-4-
hydroxyl-phenylpropanol-9-O-β-D-glucopyranoside.
References
[ 1] Jiangxi New Medical College.Dictionary of Traditional Chinese
Medicines.[ M] Beijing:China Science and Technology Pub-
lishing House.1985.2539-2540.
[ 2] Waffo-Teguo P , Hawthorne ME , Cuendet M , et al.Potential
cancer-chemopreventive activities of vine stilbenoids and fla-
vanes extracted from grape(Vitis vinifera)cell cultures [ J] .
Nutr Cancer , 2001 , 40(2):173-179.
[ 3] Zhang YW , Dou DQ , Chen YJ , et al.Effect of ginsenosides
from panax ginseng on proliferation of human osteosarcoma cell
U2OS [ J] .Chin Tradit Herb Drugs , 2001 , 32(3):232-236.
[ 4] Dou DQ , Zhang YW , Zhang L , et al.The inhibitory effects of
ginsenosides on protein tyrosine kinase activated by hypoxia
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[ J] .Planta Med , 2001 , 67(1):19-23.
[ 5] Nay B , Arnandiaud V , Vercauteren J.Gram-scale production
and applications of optically pure 13C labelled (+)-catechin
and (-)-epicatechin [ J] .Eur J Org Chem , 2001 , 2379-
2384.
[ 6] Ono M , Ito Y , Kinjo J , et al.Four new glycosides of stilbene
trimer from foeniculi fructus(fruit of foeniculum vulgare Miller)
[ J] .Chem Pharm Bull , 1995 , 43(5):868-871.
[ 7] Gamini S , Jayatilake M , Hiranthi J.Kinase inhibitors from
polygonum cuspidatum [ J] .J Natur Prod , 1993 , 56(10):
1805-1813.
[ 8] Oshima Y , Ueno Y , Hikino H , et al.Ampelopsins A , B and
C.new olicostilbenes of ampelopsis brevipedunculata var.Han-
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艹婴艹奥中分离的多酚化合物
窦德强　任 杰1　Maggie Cooper3　何曜华2　裴玉萍1　Yoshiaki Takaya2
Masatake Niwa
2　陈英杰1　姚新生1　周仁平3
(1.沈阳药科大学 天然药化教研室;2.日本Meijo 大学药学院;3.美国 Rutgers大学化学生物系)
摘要:目的　分离鉴定艹婴艹奥地上部分的化学成分 。方法　通过硅胶柱色谱 , Sephadex LH-20 及制备 HPLC 方法分离 ,对其
进行理化常数和光谱分析确定结构。结果　自艹婴艹奥地上部分分离得到 11 个多酚化合物 , 均属首次分离。结论　化合物 1 为
新化合物 ,经鉴定其结构为 3 , 5-二甲氧基-4-羟基苯丙醇-9-氧-β-D-吡喃葡萄糖苷。
关键词:艹婴艹奥;苯丙醇葡萄糖苷;多酚
59Journal of Chinese Pharmaceutical Sciences 2003 , 12(2)