Oleanolic triterpene saponins from the roots of Panax bipinnatifidus

Article abstract of DOI:10.1248/cpb.59.1417

Ten oleanane-type saponins (1-10), including three new compounds, namely bifinosides A-C (1-3), were isolated from the roots of Panax bipinnatifidus SEEM. Their structures were elucidated on the basis of chemical and spectroscopic methods.

Full text of DOI:10.1248/cpb.59.1417

November 2011
Note
Chem. Pharm. Bull. 59(11) 1417—1420 (2011)
1417
Oleanolic Triterpene Saponins from the Roots of Panax bipinnatifidus
a,b
a,b
a
b
Nguyen Huu TUNG, Tran Hong QUANG, Nguyen Thi Thanh NGAN, Chau Van MINH,
c
d
d
,a
Bui Kim ANH, Pham Quoc LONG, Nguyen Manh CUONG, and Young Ho KIM*
a
b
College of Pharmacy, Chungnam National University; Daejeon 305–764, Korea: Institute of Marine Biochemistry,
c
Vietnam Academy of Science and Technology; Institute of Chemistry, Vietnam Academy of Science and Technology; and
d
Institute of Natural Products Chemistry, Vietnam Academy of Science and Technology; 18 Hoang Quoc Viet, Caugiay,
Hanoi, Vietnam. Received June 16, 2011; accepted August 1, 2011; published online August 5, 2011
Ten oleanane-type saponins (1—10), including three new compounds, namely bifinosides A—C (1—3), were
isolated from the roots of Panax bipinnatifidus SEEM. Their structures were elucidated on the basis of chemical
and spectroscopic methods.
Key words Panax bipinnatifidus; Araliaceae; oleanane triterpene; oleanolic acid; bifinoside
Panax bipinnatifidus SEEM. (Araliaceae) is a hygrophilous
and shade-enduring plant, preferring cool and wet climate
conditions with an average temperature of about 12—15 °C.
In nature, this plant is relatively rare and mostly found in the
high mountainous region of Hoang Lien Son in the northwest
of Vietnam. The root of P. bipinnatifidus has been used as a
valuable tonic to increase mental and physical performance,
improve thinking and memory, reduce cancer risk, and lower
blood sugar in diabetics in the Vietnamese traditional medi-
1)
cine. To date, phytochemical studies on this plant have been
not undertaken extensively. In our ongoing research on
Panax plants, the presence of saponins in polar fractions of a
methanol extract of this plant was suggested. From other
plants of the same genus such as P. ginseng and P. stipu-
2—5)
leanatus, many kinds of saponins have been identified.
We therefore carried out a study of saponins of the title plant.
This paper herein deals with the isolation and structural iden-
tification of ten saponins (1—10) from the root of P. bipin-
natifidus.
Results and Discussion
The methanolic extract from the roots of P. bipinnatifidus
was suspended in water and partitioned with CH Cl . The
2
2
H O layer was subjected to Diaion HP-20 column chro-
2
matography, followed by various silica gel and C-18 re-
versed-phase columns to afford three new saponins, bifino- Fig. 1. Structures of Saponins 1—10
sides A—C (1—3), and seven known compounds including
6
)
5)
narcissiflorine methyl ester (4), chikusetsusaponin IVa (5),
feature was the signal at d 5.16 (1H, t, Jꢁ5.4 Hz), typical of
5)
12
pseudoginsenoside RP methyl ester (6), stipuleanoside R
H-12 of D oleanene skeleton, which was further confirmed
1
1
7)
5)
(
7), pseudoginsenoside RT methyl ester (8), momordin by the presence of the signals at d 123.7 (C-12) and 145.2
1
8) 5) 13 7,9)
IIe (9), and stipuleanoside R methyl ester (10), respec- (C-13) in the C-NMR spectrum. A signal at d 181.9 and
2
1
3
tively (Fig. 1).
the carbon resonances of ring D in the C-NMR spectrum
Bifinoside A (1), a white amorphous powder, has the mo- similar to those of oleanolic acid suggested the occurrence of
lecular formula C H O as determined by high resolusion- a free carboxylic group at the C-28 position. Furthermore,
4
2
66 13
electrospray ionization-time-of-flight-mass spectrometry the glycosylated 3b-hydroxy oleanolic acid was proposed as
ꢀ
(HR-ESI-TOF-MS) experiment (Found at m/z [MꢀH]
the presence of a secondary alcoholic function from the sig-
7
1
79.4596, Calcd for C H O 779.4582). Acid hydrolysis of nal at d 3.07 (1H, dd, Jꢁ11.4, 4.2 Hz), which correlated in
4
2
67 13
liberated oleanolic acid and sugar components of D-glu- the HMQC spectrum with the downfield carbon signal at d
curonic acid and L-arabinose as confirmed by gas chromatog- 91.3.
raphy (GC) analysis. The H-NMR spectrum of the aglycon
1
1
13
The H- and C-NMR spectra of 1 showed the presence of
moiety of 1 showed seven tertiary methyl groups at d 0.75, two anomeric protons at d 4.36 (d, Jꢁ5.2 Hz) and 4.44 (d,
0
.78, 0.86, 0.88, 0.89, 0.98, and 1.10, that correlated in the Jꢁ7.8 Hz) and carbons at d 105.6 and 106.3. Additionally,
heteronuclear multiple quantum coherence (HMQC) experi- the appearance of a methoxy proton at d 3.70 (3H, s), which
ments with the carbon signals at d 17.7, 15.9, 33.6, 24.0, correlated with the carbon signal at d 52.8 in the HMQC
1
6.5, 28.2, and 26.4, respectively (Tables 1—3). A further spectrum and with the carbon resonance at d 171.2 in the
∗
To whom correspondence should be addressed. e-mail: yhk@cnu.ac.kr
© 2011 Pharmaceutical Society of Japan

1
418
Vol. 59, No. 11
13
Table 1.
C-NMR Data (d, CD OD) for Bifinosides A—C (1—3)
3
Carbon
1
2
3
Carbon
1
2
3
Aglycone
Sugar moiety
3-GluAMe (I)
105.6
82.7
3-GluAMe (I)
105.8
74.7
3-GluAMe (I)
105.8
79.6
1
2
3
4
5
6
7
8
9
39.8
27.1
91.3
40.3
57.1
19.3
33.8
40.6
48.8
37.9
24.5
123.7
145.2
42.9
28.9
24.1
47.7
42.8
47.3
31.6
34.9
34.0
28.2
16.5
15.9
17.7
26.4
181.9
33.6
24.0
39.8
27.1
91.9
40.4
57.1
19.3
33.8
40.6
48.8
37.9
24.5
123.6
145.2
42.9
28.9
24.1
47.7
42.8
47.3
31.6
34.9
34.0
28.2
16.6
16.0
17.7
26.4
181.9
33.6
24.0
39.9
27.1
91.8
40.7
57.1
19.3
33.1
40.4
49.8
37.9
24.6
123.8
144.8
42.9
28.9
24.0
48.0
42.6
47.2
31.4
34.1
33.1
28.3
16.0
16.6
17.7
26.5
178.0
33.5
23.9
1
2
3
4
5
6
77.6
71.2
77.8
171.2
52.8
Ara(p) (II)
106.3
72.9
86.5
70.0
77.9
171.0
52.9
Glc (II)
105.0
74.7
78.3
71.6
79.6
67.9
Xyl (III)
104.5
76.1
78.1
71.5
66.9
86.5
71.6
76.1
170.9
52.9
Xyl (II)
104.5
76.1
78.2
71.1
COOCH3
1
2
3
4
5
6
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
74.8
67.2
64.3
66.9
Ara(p) (III)
104.9
72.9
1
2
3
4
5
74.6
67.9
64.9
28-Glc (IV)
95.6
1
2
3
4
5
6
73.9
78.6
71.4
78.2
62.4
1
1
Table 2. H-NMR Data (d, CD OD) of the Aglycon Moieties of Bifino-
Table 3. H-NMR Data (d, CD OD) of the Sugar Moieties of Bifinosides
3
3
a)
a)
sides A—C (1—3)
A—C (1—3)
H
1
2
3
H
1
2
3
b)
b)
b)
b)
b)
b)
1
0.93, 1.56, 2H
1.60, 1.77, 2H
0.93, 1.57, 2H
1.61, 1.77, 2H
0.92, 1.57, 2H
1.62, 1.76, 2H
GluAMe (I)
GluAMe (I)
GluAMe (I)
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
4.44, 1H, d (7.8) 4.46, 1H, d (7.8)
4.46, 1H, d (7.2)
3.64, 1H, t (7.8)
b)
b)
b)
b)
b)
b)
b)
b)
3.07, 1H, dd (11.4, 4.2) 3.08, 1H, dd (11.4, 4.2) 3.10, 1H, dd (11.4, 4.2)
3.56, 1H
3.48, 1H
3.42, 1H
3.81, 1H
3.48, 1H
3.69, 1H
3.78, 1H
3.83, 1H
b)
3.71, 1H
b)
0.78, 1H, dd (12.0, 4.2) 0.78, 1H, dd (12.0, 4.2) 0.78, 1H, dd (12.0, 4.2)
3.58, 1H
b)
b)
b)
b)
b)
b)
b)
1.36, 1.53, 2H
1.15, 1.26, 2H
1.36, 1.54, 2H
1.15, 1.26, 2H
1.36, 1.54, 2H
1.15, 1.26, 2H
3.85, 1H
COOCH3 3.70, 3H, s
Ara(p) (II)
3.70 s
Glc (II)
3.72, 3H, s
Xyl (II)
b)
b)
b)
1.54, 1H
1.54, 1H
1.53, 1H
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
1
2
3
4
5
6
4.36, 1H, d (5.2) 4.70, 1H, d (7.8)
4.75, 1H, d (7.2)
b)
b)
b)
b)
b)
b)
b)
b)
b)
b)
b)
b)
1 1.48, 1.88, 2H
1.48, 1.88, 2H
1.48, 1.88, 2H
3.50, 1H
3.42, 1H
3.71, 1H
3.26, 1H
3.30, 1H
3.34, 1H
3.62, 1H
3.60, 1H
3.10, 1H
b)
2 5.16, 1H, t (5.4)
5.17, 1H, t (5.4)
5.18, 1H, t (5.4)
3.39, 1H
b)
3
4
3.42, 1H
b)
b)
3.34, 3.75, 2H
3.60, 3.87, 2H
b)
b)
b)
5 1.02, 1.72, 2H
1.02, 1.72, 2H
1.04, 1.74, 2H
b)
b)
b)
6 1.80—1.91, 2H
7
1.80—1.90, 2H
1.80—1.91, 2H
3.85, 1H, dd(11.4, 5.4)
Xyl (III)
Ara(p) (III)
8 2.82, 1H, dd (12.6, 4.2) 2.80, 1H, dd (12.6, 4.2) 2.82, 1H, dd (12.6, 4.2)
1
2
3
4
5
4.41, 1H, d (7.2)
4.52, 1H, d (4.8)
b)
b)
b)
b)
b)
9 1.10, 1.65, 2H
0
1.10, 1.65, 2H
1.12, 1.67, 2H
3.12, 1H
3.56, 1H
b)
b)
3.24, 1H
3.48, 1H
b)
b)
b)
b)
b)
b)
b)
b)
1 1.17, 1.36, 2H
2 1.49, 1.71, 2H
3 0.98, 3H, s
4 0.89, 3H, s
5 0.78, 3H, s
6 0.75, 3H, s
7 1.10, 3H, s
8
1.17, 1.36, 2H
1.49, 1.71, 2H
0.98, 3H, s
0.88, 3H, s
0.77, 3H, s
0.74, 3H, s
1.10, 3H, s
1.18, 1.36, 2H
1.50, 1.71, 2H
0.99, 3H, s
0.89, 3H, s
0.78, 3H, s
0.73, 3H, s
1.10, 3H, s
3.40, 1H
3.78, 1H
b)
b)
3.13, 3.72, 2H
3.12, 3.75, 2H
Glc (IV)
1
2
3
4
5
6
5.34, 1H, d (7.8)
b)
3.29, 1H
b)
3.32, 1H
b)
3.40, 1H
b)
3.42, 1H
b)
9 0.86, 3H, s
0 0.88, 3H, s
0.85, 3H, s
0.88, 3H, s
0.86, 3H, s
0.88, 3H, s
3.63, 1H
3.80, 1H, d (12.0, 5.4)
a) Assignments were established by HMQC, DQF-COSY, and HMBC spectra. J val-
a) Assignments were established by HMQC, DQF-COSY, and HMBC spectra. J val-
ues (in Hz) are given in parentheses. b) Multiplicities not assigned due to overlapped
ues (in Hz) are given in parentheses. b) Multiplicities not assigned due to overlapped
signals.
signals.

November 2011
1419
heteronuclear multiple bond correlation (HMBC) spectrum, copyranosyloleanolic acid 3-O-b-D-xylopyranosyl(1→2)-[a-
suggested the presence of glucuronate methyl ester (Glu- L-arabinopyranosyl(1→3)]-b-D-glucuronopyranoside-6-O-
AMe) moiety. Complete assignments of all the proton reso- methyl ester.
nances in each sugar unit were achieved by double-quantum
To the best of our knowledge, there has very limited report
filtered correlation spectroscopy (DQF-COSY), HMQC, and on chemical constituents of P. bipinnatifidus. The current
HMBC spectra (Tables 1—3). In the HMBC experiment, the study has taken well part in research of phytochemical com-
anomeric signal at d 4.44 (H-1GluAMe(I)) showed a long-range ponents of the title plant, which is rich in various oleanane-
correlation with the signal at d 91.3 (C-3), indicating that type saponins. In the view of Panax genus, P. ginseng (Asian
glucuronyl moiety is directly linked to the triterpene struc- ginseng), P. quinquefolium (American ginseng), and P. viet-
ture at C-3. Subsequently, the arabinose unit linked to the namensis among the most used medicinal plants have been
GluAMe segment was at H-2GluAMe(I) on the basis of the well documented with majority of dammarane-type glyco-
1
0—14)
HMBC spectrum, in which the two anomeric protons at d sides.
Besides, the other species, P. stipuleanatus
5)
4
.36 (H-1 ) and 4.44 (H-1 ) correlated with the was dominated by oleanane-type saponins. This result can
Ara(II)
GluAMe(I)
same carbon C-2GluAMe(I) at d 82.7. Consequently, bifinoside have a significant chemotaxonomical meaning in the further
A (1) was characterized as oleanolic acid 3-O-a-L-ara- studies for classification of the Panax species.
binopyranosyl(1→2)-b-D-glucuronopyranoside-6-O-methyl
ester.
Experimental
General Procedures Optical rotations were obtained using a DIP-360
Bifinoside B (2), also a white amorphous power, revealed a digital polarimeter (Jasco, Easton, U.S.A.). IR spectra were measured using
ꢀ
a Perkin-Elmer 577 spectrometer (Perkin Elmer, Waltham, U.S.A.). NMR
spectra were recorded on Bruker DRX 600 NMR spectrometers (Bruker,
quasi-molecular ion peak [MꢀNa] at m/z 963.4913 in its
HR-ESI-TOF-MS spectrum. Acid hydrolysis of 2 afforded
Billerica, U.S.A.). HR-ESI-TOFMS experiments utilized a JEOL AccuT-
oleanolic acid and sugar components of D-glucuronic acid, D-
TM
OF LC mass spectrometer (Jeol, Tokyo, Japan). GC (Shimadzu-2010,
1
13
glucose, and D-xylose. The H- and C-NMR spectra showed
Kyoto, Japan) using a DB-05 capillary column (0.5 mm i.d.ꢂ30 m) [column
the presence of oleanolic acid, the same aglycon moiety as temperature, 210 °C; detector temperature, 300 °C; injector temperature,
2
70 °C; He gas flow rate, 30 ml/min (splitting ratio: 1/20)] was used for
saponin 1, which was glycosylated at C-3 as the downfield
shift at d 91.9, and the occurrence of three anomeric signals
sugar determination. Column chromatography (CC) was performed on silica
gel (70—230, 230—400 mesh, Darmstadt, Germany), YMC RP-18 resins
(30—50 mm, Fuji Silysia Chemical Ltd., Aichi, Japan), and HP-20 Diaion
[d 4.41
(d, Jꢁ7.2Hz), 4.70
(d, Jꢁ7.8Hz), 4.46
H
Xyl(III)
Glc(II) GluAMe(I)
(
d, Jꢁ7.8 Hz), and d 104.5, 105.0, 105.8, respectively]. The (Mitshubishi Chemical, Tokyo, Japan). TLC was performed on Kieselgel 60
C
sugar sequence was proposed by HMBC spectrum after as-
signments of the protons and the carbons by a combination
of HMQC and COSY data, starting from the anomeric pro-
ton of each sugar unit (Tables 1—3). Accordingly, H-
F
254 (1.05715; Merck) or RP-18 F254s (Merck) plates. Spots were visualized
by spraying with 10% aqueous H SO solution, followed by heating.
2
4
Plant Material The roots of P. bipinnatifidus were collected in Sapa,
Laocai, Vietnam and were taxonomically identified by botanist Ngo Van Trai
(Institute of Medicinal Materials, Hanoi, Vietnam). A voucher specimen
1GluAMe(I) (d 4.46) gave a correlation with the C-3 at d 91.9, (VHKC-0370) was deposited at the Herbarium of INPC, Vietnam.
and a cross peak between H-1
(d 4.70) and C-3
Extraction and Isolation The air-dried roots of P. bipinnatifidus
0.5 kg) were extracted in MeOH, using Soxhlet extraction apparatus (2 l,
2 h), and extracts were concentrated in vacuo to dryness. The obtained
Glc(II)
GluAMe(I)
(
1
(
d 86.5) was revealed. Additionally, a cross peak between
H-1
(d 4.41) and C-6
(d 67.9) was also observed.
Xyl(III)
Glc(II)
MeOH residue (85 g) was suspended in H O (0.5 l), then partitioned with
2
Hence, bifinoside B (2) was concluded to be oleanolic acid
CH Cl (0.5 lꢂ3) to obtain CH Cl -soluble fraction (10 g) and the water
2
2
2
2
3
-O-[b-D-xylopyranosyl(1→6)-b-D-glucopyranosyl](1→3)- layer, which was subjected to a Diaion HP-20 column eluted with a gradient
of MeOH in H O (25, 50, 75, 100% MeOH, v/v) to give four fractions (fr.
b-D-glucuronopyranoside-6-O-methyl ester.
2
1
.1—1.4). Next, fr. 1.3 (23 g) was fractionated on a silica gel column with a
Bifinoside C (3), again a white amorphous power, showed
gradient of CH Cl –MeOH (10 : 1—1 : 1) to furnish five fractions (fr. 2.1—
ꢀ
2
2
a quasi-molecular ion peak [MꢀH] at m/z 1073.5554 in the
HR-ESI-TOF-MS spectrum. The acid hydrolysis of this
2.5). Fr. 2.1 (600 mg) was repeatedly chromatographed on a silica gel col-
umn with CHCl –MeOH–H O (6 : 1 : 0.1), followed by reversed-phase
3
2
saponin liberated oleanolic acid and sugar components of D- columns eluted with MeOH–H O (4 : 1, 5 : 1) to obtain compounds 1
2
(
5.4 mg), 4 (3.5 mg), 5 (10.0 mg), and 6 (13.0 mg), respectively.
Fr. 2.2 (2.3 g) and fr. 2.3 (2.4 g) were combined due to their similar TLC
glucuronic acid, L-arabinose, D-glucose, and D-xylose based
on results of the GC analysis. Four anomeric signals were ob-
served in the C-NMR spectrum at d 95.6, 104.5, 104.9, and
1
profiles, and then subjected to a silica gel column with CHCl –MeOH–H O
(
1
3
3
2
4 : 1 : 0.1) to furnish eleven sub-fractions (fr. 3.1—3.11). Next, fr. 3.2
05.8 and at d 4.46 (1H, d, Jꢁ7.2 Hz), 4.52 (1H, d, (140 mg) was purified on a RP column with MeOH–H O (5 : 1) to yield
2
Jꢁ4.8 Hz), 4.75 (1H, d, Jꢁ7.2 Hz), and 5.34 (1H, Jꢁ7.8 Hz) compounds 2 (15.0 mg) and 7 (4.5 mg). Similarly, fr. 3.4 (220 mg) was chro-
1
matographed over a RP column with MeOH–H O (4 : 1) to yield compound
in the H-NMR spectrum. Furthermore, DQF-COSY and
2
8
(160 mg). Fr. 3.7 (280 mg) was further purified on a RP column with
HMQC experiments allowed the sequential assignments of
resonances for each sugar, starting from the anomeric proton
MeOH–H O to afford compounds 3 (45 mg) and 9 (18 mg). Finally, com-
pound 10 (75 mg) was obtained from fr. 3.9 (250 mg) by mean of RP column
2
signals (Tables 1—3). On the other hand, the HMBC spec- chromatography with the elution of MeOH–H O (4 : 1).
2
2
0
trum showed the presence of a sugar chain at the C-3 and a
monosaccharide unit at the C-28. The sequence of the mono-
saccharide chain at C-3 was further also defined by the
HMBC spectrum, in which cross peaks of H-1GluAMe(I) (d
Bifinoside A (1): White amorphous powder; [a]
D
ꢀ8.0° (cꢁ0.4, MeOH);
ꢃ1
1
IR (KBr): n 3426, 2942, 1732, 1618, 1252, 1060 cm
;
H-NMR
max
1
3
(
CD OD, 600 MHz) and C-NMR (CD OD, 150 MHz): see Tables 1—3;
3
3
ꢀ
HR-ESI-TOF-MS: m/z 779.4596 [MꢀH] (Calcd for C H O , 779.4582).
4
2
67 13
20
Bifinoside B (2): White amorphous powder; [a]D ꢀ6.4° (cꢁ0.4, MeOH);
ꢃ1
1
4
.46)/C-3 (d 91.8), H-1
(d 4.75)/C-2GluAMe(I) (d 79.6), IR (KBr): n
3424, 2940, 1731, 1618, 1250, 1059 cm
;
H-NMR
Xyl(II)
max
1
3
(
CD OD, 600 MHz) and C-NMR (CD OD, 150 MHz): see Tables 1—3;
and H-1
(d 4.52)/C-3
(d 86.5) were observed;
3
3
Ara(III)
GluAMe(I)
ꢀ
HR-ESI-TOF-MS: m/z 963.4913 [MꢀNa] (Calcd for C H O Na,
and the C-28 position of the triterpene aglycone was glycosy-
lated with the D-glucopyranosyl upon the HMBC correlation
of H-1Glc(IV) (d 5.34)/C-28 (d 178.0). On the basis of these
48 76 18
9
63.4929).
Bifinoside C (3): White amorphous powder; [a]D ꢀ18.6° (cꢁ0.7,
MeOH); IR (KBr): nmax 3428, 2946, 1718, 1624, 1248, 1054 cm ; H-NMR
20
ꢃ1
1
1
3
findings, bifinoside C (3) was identified as 28-O-b-D-glu- (CD OD, 600 MHz) and C-NMR (CD OD, 150 MHz): see Tables 1—3;
3
3

1
420
Vol. 59, No. 11
ence and Technical Publishing House, Hanoi, 2004, p. 711.
ꢀ
HR-ESI-TOFMS: m/z 1073.5554 [MꢀH] (Calcd for C H O , 1073.5533).
5
3
85 22
Acid Hydrolysis and Sugar Determination of Bifinosides A—C (1—3)
A solution of each (3.0 mg) in 2 M HCl (5.0 ml) was heated under reflux for
2) Tung N. H., Song G. Y., Park Y. J., Kim Y. H., Chem. Pharm. Bull., 57,
1412—1414 (2009).
4
h. Then, the reaction mixture was concentrated in vacuo to dryness. The
3) Tung N. H., Song G. Y., Kim J. A., Hyun J. H., Kang H. K., Kim Y. H.,
Bioorg. Med. Chem. Lett., 20, 309—314 (2010).
residue was extracted with CH Cl and H O (5 mlꢂ3). The organic portion
2
2
2
was concentrated and examined on TLC [Silica gel plates with the mobile
4) Tung N. H., Song G. Y., Minh C. V., Kiem P. V., Jin L. G., Boo H. J.,
Kang H. K., Kim Y. H., Chem. Pharm. Bull., 58, 1111—1115 (2010).
5) Liang C., Ding Y., Nguyen H. T., Kim J. A., Boo H. J., Kang H. K.,
Nguyen M. C., Kim Y. H., Bioorg. Med. Chem. Lett., 20, 7110—7115
(2010).
phase systems of CHCl –EtOAc (7 : 1) and hexane–acetone (2 : 1) compar-
3
ing with oleanolic acid (Rf 0.52, 0.45)]. Next, the sugar residue, obtained by
concentration of the water layer, was examined on TLC [Merck cellulose
plate with the mobile phase system of benzene/n-butanol/pyridine/water
(
1 : 5 : 3 : 3)] and GC (as tetramethylsilane (TMS) ethers according to the re-
6) Mei H., Kazunori O., Yutaka S., Pei-Gen X., Phytochemistry, 39,
179—184 (1995).
7) Sakai S., Katsumata M., Satoh Y., Nagasao M., Miyakoshi M., Ida Y.,
Shoji J., Phytochemistry, 35, 1319—1324 (1994).
8) Kawamura N., Watanabe H., Oshiro H., Phytochemistry, 27, 3585—
3591 (1988).
2,4)
ported method). TLC: Rf 0.22 (GluA) observed for all three hydrolysates
and the others: 0.34 (Ara) for 1; 0.30 (Glc) and 0.38 (Xyl) for 2; 0.30 (Glc),
0
(
1
.34 (Ara), and 0.38 (Xyl) for 3 in comparison with authentic sugars
Sigma-Aldrich). GC: tR (min): 4.46 (L-arabinose) for 1; 8.21 (D-xylose) and
4.12 (D-glucose) for 2; 4.46 (L-arabinose), 8.21 (D-xylose), and 14.12 (D-
glucose) for 3, which were in consistent with tR data of the standard sugars
after being treated in the similar manner [4.46 (L-arabinose), 4.90 (D-arabi-
nose), 8.21 (D-xylose), 8.66 (L-xylose), 14.12 (D-glucose), 14.25 (L-glu-
cose)].
9) Ukiya M., Akihisa T., Yasukawa K., Tokuda H., Suzuki T., Kimura Y.,
J. Nat. Prod., 69, 1692—1696 (2006).
10) Park J. D., Rhee D. K., Lee Y. H., Phytochem. Rev., 4, 159—175
(2005).
1
1) Shibata S., J. Korean Med. Sci., 16, 28—37 (2001).
Acknowledgments This study was supported by Priority Research Cen- 12) Yoshikawa M., Sugimoto S., Nakamura S., Sakumae H., Matsuda H.,
ters Program through the National Research Foundation of Korea (NRF)
Chem. Pharm. Bull., 55, 1034—1038 (2007).
funded by the Ministry of Education, Science and Technology (2009- 13) Yoshikawa M., Sugimoto S., Nakamura S., Matsuda H., Chem. Pharm.
0093815), Republic of Korea, and by Vietnam Ministry of Science and
Bull., 55, 571—576 (2007).
Technology (Project 52/2011/HD-NDT). The authors would like to thank the
KBSI for NMR measurements.
14) Duc N. M., Kasai R., Ohtani K., Ito A., Nham N. T., Yamasaki K.,
Tanaka O., Chem. Pharm. Bull., 42, 115—122 (1994).
References
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