Eranthisaponins A and B, two new bisdesmosidic triterpene saponins from the tubers of Eranthis cilicica

Article abstract of DOI:10.1021/np030071m

The present investigation aimed at the glycoside constituents of the tubers of Eranthis cilicica has resulted in the isolation of two new bisdesmosidic triterpene saponins based upon hederagenin, named eranthisaponins A (1) and B (2), along with four known triterpene saponins. The structures of the new saponins 1 and 2 were determined on the basis of spectroscopic analysis, including extensive 1D and 2D NMR data, and acid hydrolysis followed by chromatographic analysis. This is the first report concerning the secondary metabolites of E. cilicica.

Full text of DOI:10.1021/np030071m

J . Nat. Prod. 2003, 66, 879-882
879
Er a n th isa p on in s A a n d B, Tw o New Bisd esm osid ic Tr iter p en e Sa p on in s fr om
th e Tu ber s of Er a n th is cilicica
Kazuki Watanabe, Yoshihiro Mimaki,* Chiseko Sakuma, and Yutaka Sashida
School of Pharmacy, Tokyo University of Pharmacy and Life Science, 1432-1, Horinouchi, Hachioji, Tokyo 192-0392, J apan
Received February 12, 2003
The present investigation aimed at the glycoside constituents of the tubers of Eranthis cilicica has resulted
in the isolation of two new bisdesmosidic triterpene saponins based upon hederagenin, named eranthi-
saponins A (1) and B (2), along with four known triterpene saponins. The structures of the new saponins
1
and 2 were determined on the basis of spectroscopic analysis, including extensive 1D and 2D NMR
data, and acid hydrolysis followed by chromatographic analysis. This is the first report concerning the
secondary metabolites of E. cilicica.
The genus Eranthis belonging to the family Ranuncu-
laceae is composed of two species, Eranthis hyemalis and
E. cilicica, and is taxonomically related to the genus
1
Helleborus. The Eranthis plants, as well as some Helle-
borus species such as Helleborus niger and H. orientalis,
are often described as poisonous plants due to the presence
of cardiac glycosides.2 Previous phytochemical studies
disclosed the occurrence of a variety of chromenone deriv-
atives,2 a small amount of isoquinoline alkaloid, and an
-4
5
6
N-acetylgalactosamine-specific lectin in E. hyemalis, but
failed to detect cardiac glycoside even in the cardioactive
fractions prepared from E. hyemalis tubers.3 As for Er-
anthis cilicica Schott et Kotschy, indigenous to Turkey and
Afghanistan, a literature survey concerning its secondary
metabolites showed that no chemical studies have been
carried out on the plant, which prompted us to make a
phytochemical screening of the tubers of E. cilicica. Cardiac
glycosides could not be found; however, two new bisdes-
mosidic triterpene saponins based upon hederagenin,
named eranthisaponins A (1) and B (2), along with four
known triterpene saponins, were isolated. This paper deals
with the structural determination of saponins 1 and 2 on
the basis of spectroscopic analysis, including extensive 1D
and 2D NMR data, and acid hydrolysis followed by chro-
matographic analysis.
The dry tubers of E. cilicica (1.3 kg) were extracted with
hot MeOH, and the MeOH extract was passed through a
porous-polymer resin (Diaion HP-20) column. The 80%
MeOH eluate fraction was subjected to column chroma-
tography over silica gel and octadecylsilanized (ODS) silica
gel, as well as preparative HPLC, giving eranthisaponins
A (1; 15.2 mg) and B (2; 20.0 mg), along with four known
saponins.
Eranthisaponin A (1) was obtained as an amorphous
solid, and its molecular formula C71
116
H O36 was derived
from a combination of the positive-ion FABMS (m/z 1567
+
13
[
M + Na] ), C NMR spectral (71 carbon signals), and
elemental analysis data. The IR spectrum of 1 showed a
broad absorption band for hydroxyl groups at 3377 cm
-
1
,
at δ 6.21, 6.19, 5.86, 5.83, 5.06, 4.99, and 4.90. The two
three-proton doublet signals at δ 1.68 and 1.53 indicated
the presence of two deoxyhexopyranosyl units in 1. Acid
as well as absorption due to an ester carbonyl group at 1731
-1
1
cm . The H NMR spectrum of 1 was typical of a triterpene
saponin based upon an oleanolic acid derivative, showing
signals for six tertiary methyl groups at δ 1.17, 1.10, 1.07,
hydrolysis of 1 with 1 M HCl in dioxane-H
2
O (1:1) gave
3
â,23-dihydroxyolean-12-en-28-oic acid (hederagenin), to-
0
.93, 0.86, and 0.85, and an olefinic group at δ 5.38. In
gether with D-allose, L-arabinose, D-glucose, and L-rham-
nose. The monosaccharides, including their absolute con-
figurations, were identified by direct HPLC analysis of the
hydrolysate, which was performed on an aminopropyl-
addition, signals for seven anomeric protons were observed
*
To whom correspondence should be addressed. Tel: +81-426-76-4577.
Fax: +81-426-76-4579. E-mail: mimakiy@ps.toyaku.ac.jp.
1
0.1021/np030071m CCC: $25.00
© 2003 American Chemical Society and American Society of Pharmacognosy
Published on Web 05/28/2003

8
80 J ournal of Natural Products, 2003, Vol. 66, No. 6
Notes
Ta ble 1. 1H and 13C NMR Spectral Data (δ) for the Glycoside Moieties of 2 in C5D5N at 318 K
position
1H
J (Hz)
13C
position
1H
J (Hz)
13C
Ara
1
2
3
4
4.99 d
6.4
8.3, 6.4
8.3, 4.4
104.2
76.5
74.3
79.8
64.9
Rha′
Glc′′′
1
2
3
4
5
6
1
2
3
4
5
6a
b
1
2
3
4
5
6a
b
1
2
3
4
5
6
5.76 br s
4.61 br d
4.60 dd
4.40 t-like
4.90 dq
1.78 d
102.0
72.2
72.5
84.4
68.5
18.5
106.0
76.1
78.5
71.5
77.1
70.1
4.45 dd
4.09 dd
4.19 br s
4.39 br d
3.68 br d
6.12 br s
4.66 br d
4.57 dd
4.23 t-like
4.63 dq
1.63 d
3.9
9.4, 3.9
9.4
9.4, 6.1
6.1
7.8
9.1, 7.8
9.7, 9.1
9.7, 9.1
5
a
b
11.6
11.6
Rha
1
2
3
4
5
6
101.7
72.1
72.5
74.1
69.7
18.5
5.17 d
3.9
9.3, 3.9
9.3
9.3, 6.1
6.1
3.99 dd
4.12 dd
4.10 dd
3.85 m
4.60 br d
4.23 dd
4.96 d
3.91 dd
4.10 dd
4.32 t-like
3.64 m
10.5
10.5, 4.9
7.8
9.2, 7.8
9.2, 8.7
8.7
Glc
1
2
3
4
5
6
5.07 d
7.8
106.4
75.5
78.5
71.4
78.6
62.6
Glc′′′′
Rha′′
105.1
75.4
76.5
78.5
77.1
61.4
3.98 dd
4.19 dd
4.16 dd
3.85 m
4.44 br d
4.33 dd
6.19 d
4.10 dd
4.19 t-like
4.27 dd
4.07 m
4.62 br d
4.30 dd
4.95 d
3.88 dd
4.09 dd
4.32 dd
3.62 m
4.18 m
4.05 m
8.4, 7.8
9.2, 8.4
9.2, 8.5
a
b
11.0
11.0, 4.4
8.1
8.9, 8.1
8.9
8.9, 8.3
4.18 m
4.05 m
Glc′
Glc′′
1
2
3
4
5
6
95.7
74.0
78.7
71.0
77.9
69.4
5.77 br s
4.64 br d
4.49 dd
4.27 t-like
4.86 dq
1.67 d
102.6
72.5
72.8
73.9
70.3
18.5
3.8
9.1, 3.8
9.1
9.1, 6.1
6.1
a
b
10.9
10.9, 4.9
7.8
8.9, 7.8
8.9, 8.4
9.4, 8.4
1
2
3
4
5
6
104.9
75.4
76.5
77.9
77.1
61.5
a
b
bonded silica gel column using MeCN-H
solvent system, with detection being carried out by using
2
O (17:3) as
28-O-R-L-rhamnopyranosyl-(1f4)-O-â-D-glucopyranosyl-
(1f6)-â-D-glucopyranoside, which was assigned to 1.
Eranthisaponin B (2) was isolated as an amorphous solid
1
an optical rotation (OR) detector. Comparison of the H and
1
3
C NMR spectra of 1 with those of 3â-[(O-â-D-gluco-
with a molecular formula C83
data of the positive-ion FABMS (m/z 1875 [M + Na] ),
136
H O45, as determined from
pyranosyl-(1f4)-O-[R-L-rhamnopyranosyl-(1f2)]-R-L-arabino-
pyranosyl)oxy]-23-hydroxyolean-12-en-28-oic acid 28-O-R-
L-rhamnopyranosyl-(1f4)-O-â-D-glucopyranosyl-(1f6)-â-D-
+
13
C
NMR spectrum (83 carbon signals), and elemental analysis.
1
The H NMR spectrum of 2 showed signals for nine
7
glucopyranoside (3) revealed that the aglycon whose C-3
anomeric protons at δ 6.19, 6.12, 5.77, 5.76, 5.17, 5.07, 4.99,
hydroxyl and C-28 carboxyl groups were substituted by the
sugar moieties and the triglycoside residue attached at
C-28 of the aglycon were identical to those of 3. However,
significant differences were recognized in the signals from
the rhamnosyl moiety linked to the inner arabinosyl group
and the presence of the additional six carbon signals of a
hexose at δ 104.2 (CH), 75.9 (CH), 73.0 (CH), 72.7 (CH),
4
.96, and 4.95, along with signals for six tertiary methyl
groups at δ 1.18, 1.11, 1.06, 0.98, 0.89, and 0.87, and an
olefinic group at δ 5.40 as observed for 1. The three methyl
doublet signals at δ 1.78, 1.67, and 1.63 were indicative of
the presence of three 6-deoxyhexosyl units in 2. Acid
hydrolysis of 2 yielded hederagenin, L-arabinose, D-glucose,
and L-rhamnose. The 13C NMR shifts of C-3 at δ 81.1 and
C-28 at δ 176.5 implied that the sugar linkages were at
both C-3 and C-28. From the above data, 2 was assumed
to be a hederagenin 3,28-bisdesmoside whose sugar moi-
eties were composed of a total of nine monosaccharides.
The severe overlapping of the proton signals for the sugar
moieties excluded the possibility of a complete assignment
in a straightforward way using the conventional 2D NMR
6
2
8.9 (CH), and 62.8 (CH ). The anomeric carbon signal at
δ 104.2 was associated with the anomeric proton signal at
δ 5.86 with a relatively large J value of 7.9 Hz by the
HMQC spectrum. Analysis of the 1H- H COSY and 2D
TOCSY spectra, starting from the δ 5.86 anomeric reso-
nance, allowed the sequential assignments of the signals
1
2
from H-1 to CH -6 including their multiplet patterns and
1
1
coupling constants, which were indicative of a â-D-allo-
methods such as the H- H COSY, 2D TOCSY, and HSQC
spectra in 2. Analysis of the 1D TOCSY spectra followed
by interpretation of the 1H- H COSY, HSQC, HSQC-
TOCSY, and HMBC spectra allowed us to solve the exact
sugar sequences of the sugar moieties and their linkage
8
pyranosyl group. In the HMBC spectrum, the anomeric
1
proton of the allosyl moiety showed a long-range correlation
with the δ 82.8 signal, assignable to C-3 of the rhamnosyl
unit, of which the anomeric proton at δ 6.19, in turn,
3
1
showed a J C,H correlation with C-2 of the arabinosyl at δ
positions to the aglycon. The H NMR subspectra of
7
5.8. An HMBC correlation from the anomeric proton of
individual monosaccharide units were obtained by using
selective irradiation of the easily identifiable anomeric
proton signals, as well as irradiation of other nonover-
lapping proton signals in a series of 1D TOCSY experi-
ments. Subsequent analysis of the H- H COSY spectrum
resulted in the sequential assignments of all the proton
the terminal glucosyl at δ 5.06 to C-4 of the arabinosyl at
δ 80.5 was also noted. All of these data were consistent
with the structure 3â-[(O-â-D-allopyranosyl-(1f3)-O-R-L-
rhamnopyranosyl-(1f2)-O-[â-D-glucopyranosyl-(1f4)]-R-L-
arabinopyranosyl)oxy]-23-hydroxyolean-12-en-28-oic acid
9
1
1

Notes
J ournal of Natural Products, 2003, Vol. 66, No. 6 881
resonances due to the nine monosaccharides, including
identification of their multiplet patterns and coupling
constants as shown in Table 1. The HSQC and HSQC-
TOCSY spectra correlated the proton resonances with those
of the corresponding one-bond coupled carbons, leading to
unambiguous assignments of the carbon shifts (Table 1).
and Diaion HP-20 (Mitsubishi-Chemical, Tokyo, J apan) were
used for column chromatography. TLC was carried out on
precoated Kieselgel 60 F254 (0.25 mm thick, Merck, Darmstadt,
Germany) and RP-18 F254S (0.25 mm thick, Merck) plates, and
2 4
spots were visualized by spraying the plates with 10% H SO
solution, followed by heating. HPLC was performed using a
system comprised of a Tosoh CCPM pump (Tokyo, J apan), a
Tosoh CCP PX-8010 controller, a Tosoh RI-8010 detector, a
Shodex OR-2 detector (Showa-Denko, Tokyo, J apan), and a
Rheodyne injection port. Capcell Pak C18 UG80 columns (10
mm i.d. × 250 mm, 5 µm, Shiseido, Tokyo, J apan) were
employed for preparative HPLC.
P la n t Ma ter ia l. Eranthis cilicica was purchased from a
nursery in Heiwaen, J apan, in October 2000, and was identi-
fied by one of the authors (Y.S.). A voucher specimen has been
deposited in our laboratory (voucher No. 00-7-EC, Laboratory
of Medicinal Plant Science).
Comparison of the carbon chemical shift thus assigned with
those of the reference methyl glycosides,10,11 taking into
account the known effects of O-glycosylation, indicated that
2
contained a â-D-glucopyranosyl unit (Glc) and two R-L-
rhamnopyranosyl units (Rha and Rha′′) as the terminal
glycosyl moieties, and an R-L-arabinopyranosyl unit (Ara),
four â-D-glucopyranosyl units (Glc′, Glc′′, Glc′′′, and Glc′′′′),
and an R-L-rhamnopyranosyl unit (Rha′) as the substituted
sugar moieties. The relatively large J values of the ano-
meric protons of the arabinosyl (6.4 Hz) and glucosyl (7.8-
8
.1 Hz) moieties indicated an R anomeric orientation for
Extr a ction a n d Isola tion . Dry tubers of E. cilicica (1.3
kg) were extracted with hot MeOH (20 L). Following removal
of MeOH, the residue (135 g) was passed through a Diaion
HP-20 column (200 mm i.d. × 200 mm), eluted with 30%
MeOH, 50% MeOH, 80% MeOH, MeOH, EtOH, and EtOAc.
The 80% MeOH eluate portion (30 g) was subjected to column
chromatography on silica gel (80 mm i.d. × 280 mm) and
the arabinosyl and â for the glucosyls. For the rhamnosyl
moieties, the large J C,H values (>170 Hz) confirmed that
1
the anomeric protons were equatorial (R-pyranoid anomeric
form). In the HMBC spectrum, the anomeric proton signals
at δ 6.12 (Rha) and 5.07 (Glc) showed long-range correla-
tions with C-2 at δ 76.5 and C-4 at δ 79.4 of Ara,
respectively, whose anomeric proton signal at δ 4.99, in
turn, exhibited a long-range correlation with C-3 of the
aglycon at δ 81.1, indicating that the branched triglycoside
attached at C-3 of the aglycon was the same as that of 3.
Consequently, an oligoglycoside composed of six mono-
saccharides was presumed to be linked to C-28 of the
aglycon. Further HMBC correlations from δ 5.77 (H-1 of
Rha′′) to C-4 of Glc′′′′ at δ 78.5, δ 4.96 (H-1 of Glc′′′′) to C-6
of Glc′′′ at δ 70.1, δ 5.17 (H-1 of Glc′′′) to C-4 of Rha′ at δ
eluted with stepwise gradient mixtures of CHCl -MeOH (19:
3
1
; 9:1; 4:1; 2:1) and finally with MeOH to give 38 fractions.
Fractions with the same TLC profile were combined, and nine
fractions (I-IX) were recovered. Fraction IX was further
divided by column chromatography on silica gel (65 mm i.d.
× 280 mm) eluted with CHCl
3
2
-MeOH-H O (14:8:1) into four
subfractions (IX-1-IX-4). Fraction IX-1 was subjected to an
ODS silica gel column (45 mm i.d. × 190 mm) eluted with
2
MeCN-H O (2:5) to give 23-hydroxy-3â-[(O-R-L-rhamno-
pyranosyl-(1f2)-R-L-arabinopyranosyl)oxy]olean-12-en-28-oic
acid 28-O-R-L-rhamnopyranosyl-(1f4)-O-â-D-glucopyranosyl-
8
4.4, δ 5.76 (H-1 of Rha′) to C-4 of Glc′′ at δ 77.9, δ 4.95
(
1f6)-â-D-glucopyranoside (97.8 mg).14 Fraction IX-2 was
(H-1 of Glc′′) to C-6 of Glc′ at δ 69.4, and δ 6.19 (H-1 of
subjected to ODS silica gel column chromatography (45 mm
i.d. × 190 mm) eluted with MeCN-H O (4:11) and preparative
HPLC using MeCN-H O (2:5) to yield 3â-[(O-â-D-gluco-
Glc′) to C-28 of the aglycon at δ 176.5 confirmed the
hexaglycoside sequence as Rha′′-(1f4)-Glc′′′′-(1f6)-Glc′′′-
2
2
(1f4)-Rha′-(1f4)-Glc′′-(1f6)-Glc′, which was attached at
pyranosyl-(1f4)-R-L-arabinopyranosyl)oxy]-23-hydroxyolean-
12-en-28-oic acid 28-O-R-L-rhamnopyranosyl-(1f4)-O-â-D-gluco-
pyranosyl-(1f6)-â-D-glucopyranoside (10.5 mg).15 Fraction IX-3
was chromatographed on ODS silica gel (45 mm i.d. × 190
C-28 of the aglycon. Accordingly, the structure of 2 was
determined as 3â-[(O-â-D-glucopyranosyl-(1f4)-O-[R-L-
rhamnopyranosyl-(1f2)]-R-L-arabinopyranosyl)oxy]-23-hy-
droxyolean-12-en-28-oic acid 28-O-R-L-rhamnopyranosyl-
2
mm) eluted with MeCN-H O (1:3) to give 3 (77.4 mg) and a
mixture of two saponins, which was separated by preparative
HPLC using MeCN-H O (1:3) to furnish 1 (15.2 mg) and 3â-
(O-â-D-glucopyranosyl-(1f2)-O-[â-D-glucopyranosyl-(1f4)]-R-
(
1f4)-O-â-D-glucopyranosyl-(1f6)-O-â-D-glucopyranosyl-
2
(
(
1f4)-O-R-L-rhamnopyranosyl-(1f4)-O-â-D-glucopyranosyl-
[
1f6)-â-D-glucopyranoside.
L-arabinopyranosyl)oxy]-23-hydroxyolean-12-en-28-oic acid 28-
O-R-L-rhamnopyranosyl-(1f4)-O-â-D-glucopyranosyl-(1f6)-â-
D-glucopyranoside (42.5 mg).15 Compound 2 (20.0 mg) was
isolated from fraction IX-4 by subjecting it to preparative
This is the first report concerning the secondary me-
tabolites of E. cilicica, and eranthisaponins A (1) and B
2) are newly described bisdesmosidic triterpene saponins.
(
HPLC using MeCN-H
Er a n th isa p on in A (1): amorphous solid; [R]
.10, MeOH); IR (film) νmax 3377 (OH), 2926 (C-H), 1731
2
O (1:3).
The branched tetraglycoside attached at C-3 of the aglycon
in 1 and the linear hexaglycoside attached at C-28 of the
aglycon in 2 have not been reported as sugars of either
triterpene saponins or steroidal saponins. It is also notable
that 1 has been found to contain D-allopyranose as a sugar
component, which is rarely encountered in the plant
saponins.1
2
6
D
-22.0° (c
0
-
1
(
CdO), 1453, 1387, 1365, 1257, 1232, 1075, 1035, 913 cm ;
1
H NMR (pyridine-d ) δ 6.21 (1H, d, J ) 8.1 Hz, H-1 of Glc′),
5
6
.19 (1H, br s, H-1 of Rha), 5.86 (1H, d, J ) 7.9 Hz, H-1 of
All), 5.83 (1H, br s, H-1 of Rha′), 5.38 (1H, t-like, J ) 3.2 Hz,
2,13
H-12), 5.06 (1H, d, J ) 7.8 Hz, H-1 of Glc), 4.99 (1H, d, J )
7
(
5
4
.8 Hz, H-1 of Glc′′), 4.90 (1H, d, J ) 7.1 Hz, H-1 of Ara), 4.71
Exp er im en ta l Section
1H, t-like, J ) 3.0 Hz, H-3 of All), 4.48 (1H, br dd, J ) 8.7,
.2 Hz, H-5 of All), 4.42 (1H, br d, J ) 11.7 Hz, H-6a of All),
.36 (1H, d, J ) 11.7 Hz, H-23a), 4.28 (1H, dd, J ) 11.7, 5.2
Gen er a l Exp er im en ta l P r oced u r es. Optical rotations
were measured using a J ASCO DIP-360 (Tokyo, J apan)
automatic digital polarimeter. IR spectra were recorded on a
J ASCO FT-IR 620 spectrophotometer. NMR spectra were
Hz, H-6b of All), 4.20 (1H, m, H-3), 4.18 (1H, dd, J ) 8.7, 3.0
Hz, H-4 of All), 4.01 (1H, dd, J ) 7.9, 3.0 Hz, H-2 of All), 3.57
(
1H, d, J ) 11.7 Hz, H-23b), 3.14 (1H, dd, J ) 13.5, 3.8 Hz,
1
recorded on a Bruker DRX-500 spectrometer (500 MHz for H
H-18), 1.68 (3H, d, J ) 6.2 Hz, Me-6 of Rha′), 1.53 (3H, d, J )
NMR, Karlsruhe, Germany) using standard Bruker pulse
programs. Chemical shifts are given as δ values with reference
to tetramethylsilane (TMS) as internal standard. MS were
recorded on a Finnigan MAT TSQ-700 (San J ose, CA) mass
spectrometer, using a dithiothreitol and dithioerythritol (3:1)
matrix. Silica gel (200-400 mesh, Fuji-Silysia Chemical, Aichi,
J apan), ODS silica gel (75 µm, Nacalai Tesque, Kyoto, J apan),
6
1
0
.2 Hz, Me-6 of Rha), 1.17 (3H, s, Me-27), 1.10 (3H, s, Me-24),
.07 (3H, s, Me-26), 0.93 (3H, s, Me-25), 0.86 (3H, s, Me-30),
.85 (3H, s, Me-29); C NMR (pyridine-d ) δ 39.0, 26.4, 81.1,
13
5
43.5, 47.5, 18.0, 32.7, 39.8, 48.1, 36.8, 23.8, 122.8, 144.1, 42.1,
28.2, 23.3, 47.0, 41.6, 46.1, 30.7, 33.9, 32.5, 63.7, 14.0, 16.1,
17.5, 26.0, 176.5, 33.0, 23.6 (aglycon C-1-C-30), 104.7, 75.8,
74.9, 80.5, 65.9 (Ara C-1-C-5), 101.3, 71.5, 82.8, 72.7, 69.8,

8
82 J ournal of Natural Products, 2003, Vol. 66, No. 6
Notes
1
8.5 (Rha C-1-C-6), 104.2, 73.0, 72.7, 68.9, 75.9, 62.8 (All C-1-
30.7, 34.1, 32.6, 64.1, 13.9, 16.2, 17.6, 26.1, 176.5, 33.1, 23.7
C-6), 106.7, 75.4, 78.7, 71.2, 78.6, 62.4 (Glc C-1-C-6), 95.6,
(aglycon C-1-C-30), signals for the sugar moieties, see Table
+
7
7
1
3.8, 78.4, 70.8, 78.0, 69.1 (Glc′ C-1-C-6), 104.7, 75.3, 76.5,
8.2, 77.1, 61.2 (Glc′′ C-1-C-6), 102.6, 72.5, 72.7, 73.9, 70.2,
1; FABMS (positive mode) m/z 1975 [M + Na] ; anal. C
51.40%, H 7.46%, calcd for C83
7.55%.
H
136
O
2
45‚9/2H O, C 51.52%, H
8.5 (Rha′′ C-1-C-6); FABMS (positive mode) m/z 1567 [M +
+
Na] ; anal. C 52.65%, H 7.86%, calcd for C71
H
116
O
2
36‚4H O, C
Acid Hyd r olysis of 2. Compound 2 (7.6 mg) was subjected
to acid hydrolysis as described for 1 to give hederagenin (1.2
mg) and a sugar fraction (3.5 mg). HPLC analysis of the sugar
fraction under the same conditions as in the case of that of 1
showed the presence of L-arabinose, D-glucose, and L-rham-
5
2.71%, H 7.73%.
Acid Hyd r olysis of 1. A solution of 1 (5.2 mg) in 1 M HCl
dioxane-H O, 1:1, 3 mL) was heated at 95 °C for 1 h under
an Ar atmosphere. After the reaction mixture was diluted with
(
2
H
2
O (2 mL), it was extracted with Et
extract was chromatographed on silica gel eluted with CHCl
MeOH (19:1) to give hederagenin (0.9 mg). The H
2
O (5 mL × 3). The Et
2
O
-
nose: t
R
(min) 7.09 (L-rhamnose, negative optical rotation), 8.13
3
(L-arabinose, positive optical rotation), 13.25 (D-glucose, posi-
2
O residue
tive optical rotation).
was neutralized using an Amberlite IRA-93ZU (Organo, Tokyo,
J apan) column and passed through a Sep-Pak C18 cartridge
Su p p or tin g In for m a tion Ava ila ble: Spectroscopic data (13
NMR, H- H COSY, HSQC, HMBC, HSQC-TOCSY, and 1D TOCSY
spectra) for compound 2. This material is available free of charge via
the Internet at http://pubs.acs.org.
C
(Waters, Milford, MA) eluted with H
2
O-MeOH (3:2) to give a
1
1
sugar fraction (2.6 mg). The sugar fraction was analyzed by
HPLC under the following conditions: column, Capcell Pak
NH2 UG80 (4.6 mm i.d. × 250 mm, 5 µm, Shiseido); solvent,
2
MeCN-H O (17:3); flow rate, 0.9 mL/min; detection, RI and
Refer en ces a n d Notes
optical rotation (OR). Identification of D-allose, L-arabinose,
D-glucose, and L-rhamnose was carried out by comparison of
its retention time and OR with those of an authentic sample;
(
1) Tsukamoto, Y., Exec. Ed. The Grand Dictionary of Horticulture;
Shogakukan: Tokyo, 1988; Vol. 2, pp 89-92.
(
2) Kopp, B.; Kubelka, E.; Reich, C.; Robien, W.; Kubelka, W. Helv. Chim.
Acta 1991, 74, 611-616.
t
(
R
(min): 7.09 (negative optical rotation, L-rhamnose), 8.13
positive optical rotation, L-arabinose), 10.31 (positive optical
(
(
(
3) Egger, K. Z. Naturforsch. 1961, 16b, 697-702.
4) J unior, P. Phytochemistry 1979, 18, 2053-2054.
5) Slav ´ı k, J .; Bochor a´ kov a´ , J .; Slav ´ı kov a´ , L. Collect. Czech. Chem.
Commun. 1987, 52, 804-812.
rotation, D-allose), 13.16 (positive optical rotation, D-glucose).
2
6
Er a n th isa p on in B (2): amorphous solid; [R]
D
-36.0° (c
0
1
.10, MeOH); IR (film) νmax 3375 (OH), 2922 and 2852 (C-H),
732 (CdO), 1451, 1387, 1365, 1255, 1232, 1063, 1034, 913
(6) Cammue, B. P.; Peeters, B.; Peumans, W. J . Biochem. J . 1985, 227,
49-955.
9
-
1
1
(7) Kang, S. S. Arch. Pharm. Res. 1989, 12, 42-47.
cm ; H NMR (pyridine-d
Glc′), 6.12 (1H, br s, H-1 of Rha), 5.77 (1H, br s, H-1 of Rha′′),
5
5
H-1 of Glc), 4.99 (1H, d, J ) 6.4 Hz, H-1 of Ara), 4.96 (1H, d,
J ) 7.8 Hz, H-1 of Glc′′′′), 4.95 (1H, d, J ) 7.8 Hz, H-1 of Glc′′),
4
d, J ) 10.5 Hz, H-23b), 3.16 (1H, dd, J ) 13.9, 3.9 Hz, H-18),
1
5
) δ 6.19 (1H, d, J ) 8.1 Hz, H-1 of
(
8) Mimaki, Y.; Sashida, Y.; Shimomura, H. Phytochemistry 1989, 28,
3453-3458.
.76 (1H, br s, H-1 of Rha′), 5.40 (1H, t-like, J ) 3.3 Hz, H-12),
.17 (1H, d, J ) 7.8 Hz, H-1 of Glc′′′), 5.07 (1H, d, J ) 7.8 Hz,
(9) Watanabe, K.; Mimaki, Y.; Sakuma, C.; Sashida, Y. Chem. Lett. 2002,
3
1, 772-773.
(
10) Agrawal, P. K.; J ain, D. C.; Gupta, R. K.; Thakur, R. S. Phytochemistry
1
985, 24, 2479-2496.
(11) Agrawal, P. K. Phytochemistry 1992, 31, 3307-3330.
.18 (1H, d, J ) 10.5 Hz, H-23a), 4.17 (1H, m, H-3), 3.73 (1H,
(12) Okabe, H.; Miyahara, Y.; Yamauchi, T. Tetrahedron Lett. 1982, 23,
7
7-80.
(
13) Baykal, T.; Panayir, T.; Tasdemir, D.; Sticher, O.; C¸ alis, I. Phytochem-
.78 (3H, d, J ) 6.1 Hz, Me-6 of Rha′), 1.67 (3H, d, J ) 6.1
istry 1998, 48, 867-873.
Hz, Me-6 of Rha′′), 1.63 (3H, d, J ) 6.1 Hz, Me-6 of Rha), 1.18
(14) Sano, K.; Sanada, S.; Ida, Y.; Shoji, J . Chem. Pharm. Bull. 1991, 39,
865-870.
(
(
3H, s, Me-27), 1.11 (3H, s, Me-26), 1.06 (3H, s, Me-24), 0.98
3H, s, Me-25), 0.89 (3H, s, Me-30), 0.87 (3H, s, Me-29); 13
(15) Li, X.-C.; Wang, D.-Z.; Wu, S.-G.; Yang, C.-R. Phytochemistry 1990,
C
2
9, 595-599.
NMR (pyridine-d
4
5
) δ 39.1, 26.2, 81.1, 43.5, 47.9, 18.2, 32.9, 40.0,
8.2, 36.9, 23.9, 123.0, 144.2, 42.2, 28.4, 23.5, 47.1, 41.8, 46.3,
NP030071M