Tetrapterosides A and B, two new oleanane-type saponins from Tetrapleura tetraptera

Article abstract of DOI:10.1002/mrc.2381

From the stem bark of Tetrapleura tetraptera, two new oleanane-type saponins, tetrapteroside A 3-O-{6-O-[(2E,6S)-2,6-dimethyl-6-hydroxyocta-2,7- dienoyl]-β-D-glucopyranosyl-(1 → 2)-β-D-glucopyranosyl-(1 → 3)-β-D-glucopyranosyl-(1 → 4)-[β-D-glucopyranosyl-

Full text of DOI:10.1002/mrc.2381

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Spectral Assignments and Reference Data
Received: 12 June 2008
Revised: 27 October 2008
Accepted: 31 October 2008
Published online in Wiley Interscience: 19 January 2009
(www.interscience.com) DOI 10.1002/mrc.2381
Tetrapterosides A and B, two new
oleanane-type saponins from Tetrapleura
tetraptera
a,b
a
c
´
Olivier Placide Note, Anne-Claire Mitaine-Offer, Tomofumi Miyamoto,
d
b
´
Thomas Paululat, Dieudonne Emmanuel Pegnyemb and Marie-Aleth
Lacaille-Duboisa∗
From the stem bark of Tetrapleura tetraptera, two new oleanane-type saponins, tetrapteroside A 3-O-{6-O-[(2E,6S)-
2,6-dimethyl-6-hydroxyocta-2,7-dienoyl]-β-D-glucopyranosyl-(1 → 2)-β-D-glucopyranosyl-(1 → 3)-β-D-glucopyranosyl-(1 →
4)-[β-D-glucopyranosyl-(1 → 2)]-β-D-glucopyranosyl}-3,27-dihydroxyoleanolic acid (1), and tetrapteroside B 3-O-{β-D-
glucopyranosyl-(1 → 2)-6-O-[(E)-feruloyl]-β-D-glucopyranosyl-(1 → 3)-β-D-glucopyranosyl-(1 → 4)-[β-D-glucopyranosyl-(1 →
2)]-β-D-glucopyranosyl}-3,27-dihydroxyoleanolic acid (2), were isolated. Further extractions from the roots led to the isolation
of four known oleanane-type saponins. Their structures were elucidated by the combination of mass spectrometry (MS), one
c
and two-dimensional NMR experiments. Copyright ꢀ 2009 John Wiley & Sons, Ltd.
Keywords: NMR; 1H; 13C; 2D NMR; triterpene saponins; tetrapteroside A; tetrapteroside B; Mimosaceae; Tetrapleura tetraptera
Introduction
were observed at m/z = 1281 [(M − H) −166]−, 1119 [(M − H)–166
−162]−, 957[(M−H)–166−162−162]−, and795[(M−H)–166−
162 − 162 − 162]−, corresponding to the successive loss of one
monoterpenoyl moiety, and three hexosyl moieties, respectively.
The 1H-NMR spectrum of 1 showed characteristic signals
of a saponin structure composed by an aglycon part and
an oligosaccharidic part. For the aglycon moiety, the 1H-NMR
spectrum displayed signals for six angular methyl groups as
singlets, one olefinic proton at δH 5.77 (br t, J = 3 Hz, H-12),
one oxygen bearing methine protons at δH 3.08 (dd, J = 11.4,
3.8 Hz, H-3), and one primary alcoholic function at C-27 position,
δH 3.72 (d, J = 10.5 Hz), 3.98. In the 13C-NMR spectrum, the
deshielded signal at δC 89.5 (C-3) in comparison with the free
aglycon, suggested a glycosidic linkage at C-3. The structure
of the aglycon of 1 was thus recognized to be the triterpene
3,27-dihydroxyoleanolic acid by 1H-NMR and 13C-NMR analyses
In a continuation of the study on bioactive triterpene saponins
from Cameroonian medicinal plants,[1–5] we have examined the
saponin fraction of the stem bark and roots of Tetrapleura
tetraptera Taub. (Mimosaceae), locally named Aridan. This ro-
bust tree from the forest of tropical Africa is well-known as
remedies in the traditional medicine,[6] and for its strong mol-
luscicidal activity. The previous chemical analysis of the fruit
and stem bark led to the isolation of oleanane-type triterpene
saponins like aridanin, a N-acetylglycoside triterpenoid.[7,8] In
this paper, we report the isolation by successive chromato-
graphic steps of two new triterpene saponins, tetrapterosides
A (1) and B (2) (Fig. 1) from the stem bark and four known
saponins from the roots,[7,8] 3-O-(2-acetamido-2-deoxy-β-D-gluco-
pyranosyl)oleanolic acid (aridanin), 3-O-(2-acetamido-2-deoxy-
β-D-glucopyranosyl)echinocystic acid, 3-O-[β-D-glucopyranosyl-
(1 → 6)-2-acetamido-2-deoxy-β-D-glucopyranosyl]oleanolic acid,
and 3-O-sulfoechinocystic acid. Their structures were established
mainlybytwo-dimensional(2D)NMR(COSY,TOCSY,NOESY,HSQC,
HMBC) and mass spectrometry.
∗
Correspondence to: Marie-Aleth Lacaille-Dubois, Laboratoire de Pharmacog-
´
´
nosie, UMIB, UPRES EA 3660, Faculte de Pharmacie, Universite de Bourgogne, 7
bd. Jeanne D’ Arc, BP 87900, 21079 Dijon cedex, France.
E-mail: m-a.lacaille-dubois@u-bourgogne.fr
´
Laboratoire de Pharmacognosie, UMIB, UPRES EA 3660, Faculte de Pharmacie,
a
Results and Discussion
´
Universite de Bourgogne, 7 bd. Jeanne D’ Arc, BP 87900, 21079 Dijon cedex,
France
Compound 1, a white amorphous powder, exhibited in high-
resolution electrospray ionization mass spectrometry (HR-ESIMS)
(positive-ion mode) a pseudo-molecular ion peak at m/z =
1471,7090 [M + Na]+ (calcd 1471,7085), consistent with a
molecular formula of C70H112O31Na. Its fast-atom bombardment
mass spectrum (FABMS) (negative-ion mode) showed a quasi-
molecular ion peak at m/z = 1447 [M − H]−, indicating a
molecular weight of 1448. Other significant fragment ion peaks
´ ´ ´
Departement de Chimie Organique, Faculte des Sciences, Universite de
Yaounde, BP 812 Yaounde, Cameroun
b
c
´
´
Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka
812-8582, Japan
¨
d
Universitat Siegen FB8, OC- II (AK Ihmels) Adolf-Reichwein-Str. 2 D- 57068
Siegen, Germany
c
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O. P. Note´ et al.
COOH
GlcI
GlcIII
OH
OH
CH2OH
O
3
O
O
O
HO
GlcIV
HO
OH
O
OR1
O
OH
O
HO
HO
O
HO
HO
O
OR2
OH
GlcII
O
HO
HO
OH
GlcV
R
R
2
1
O
1
9
2
1
H
3
6
4
8
MT
5
7
OH
10
OCH3
OH
FA
2
3
H
H
2
4
5
γ
α
1
β
6
OH
Figure 1. Structures of 1 and 2.
GlcI
OH
GlcIII
OH
3
O
O
O
O
HO
GlcIV
HO
HO
O
OH
O
OH
O
OH
HO
O
HO
HO
O
OMT
O
OH
GlcII
HO
HO
OH
1
GlcV
GlcI
GlcIII
OH
3
OH
O
O
O
O
O
HO
GlcIV
HO
HO
O
OFA
O
OH
OH
HO
O
HO
HO
O
OH
OH
O
GlcII
HO
HO
OH
GlcV
2
(
)
correlations for 1 and 2.
Figure 2. Important HMBC
and NOESY
)
(
c
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Spectral Assignments and Reference Data
peaks in the NOESY spectrum between δH 4.27 (GlcI-2) and an
anomeric signal at δH 5.44 (GlcII-1), and between δH 4.07 (GlcI-
4) and another anomeric signal at δH 4.95 (GlcIII-1) (Fig. 2). In
the HMBC spectrum, the correlation between δH 3.74 (GlcIII-3)
and δC 103.2 (GlcIV-1), and the reverse correlation between δH
(GlcIV-1) and δC 90.1 (GlcIII-3), and the correlation between δH
3.96 (GlcIV-2) and δC 105.7 (GlcV-1), revealed the (1 → 3) linkage
between GlcIV and GlcIII and the (1 → 2) linkage between GlcV
and Glc IV (Fig. 2). Moreover, the deshielded signal of GlcV-6 at
δC 63.9 and δH 4.71, 5.13 showed an acylation at this position.
After subtraction of the signals of the oligosaccharidic chain
linked at the C-3 position, signals of a monoterpenoyl residue
still remained, which acylated the GlcV-6 position (Table 3). Its
NMR data are in accordance with those described in literature[11]
for a (2E,6S)-2,6-dimethyl-6-hydroxyocta-2,7-dienoyl unit, already
foundinacylatedsaponinsisolatedfromplantsoftheMimosaceae
family.[11,12]
Table 1. 1H-NMR (600 MHz) and 13C-NMR(150 MHz) data of the
aglycons of 1 and 2 in pyridine-d5 (δ in ppm, J in Hz)
Compound 1
1H-NMR
Compound 2
1H-NMR
No
13C-NMR
13C-NMR
1
0.83, 1.28
38.3
26.0
89.5
39.3
55.3
18.4
33.8
40.0
48.2
36.7
23.6
127.6
139.7
47.5
24.1
23.5
46.3
41.4
45.3
30.6
33.2
32.8
27.6
16.4
15.4
18.5
64.1
179.5
32.9
23.6
0.87, 1.31
38.2
26.0
89.6
39.5
55.3
18.2
33.8
40.0
48.2
36.7
23.4
127.1
139.8
47.5
23.9
23.6
46.4
41.5
45.4
30.6
33.1
32.9
27.7
16.4
15.5
18.5
63.9
179.5
32.9
23.6
2
1.75, 1.95
1.76, 1.96
3
3.08 dd, J = 11.4, 3.8
3.12 dd, J = 11.6, 3.8
4
–
–
5
0.81
0.83
6
1.24, 1.51
1.24, 1.50
7
1.04, 1.25
1.08, nd
8
–
–
9
2.05
2.07
10
11
12
13
14
15
16
17
–
–
1.88, 2.04
1.94, 2.03
The structure of 1 was thus established as 3-O-{6-O-[(2E,6S)-2,6-
5.77 br t, J = 3.0
5.84 br t, J = 3.0
dimethyl-6-hydroxyocta-2,7-dienoyl]-β-D-glucopyranosyl-(1
→
–
–
–
2)-β-D-glucopyranosyl-(1
[β-D-glucopyranosyl-(1 → 2)]-β-D-glucopyranosyl}-3,27-dihydro-
→
3)-β-D-glucopyranosyl-(1
→
4)-
–
nd
nd
–
nd
xyoleanolic acid,
tetrapteroside A.
a new oleanane-type glycoside named
nd
–
Compound 2, a white amorphous powder, exhibited in HR-
ESIMS (positive-ion mode) a pseudo-molecular ion peak at m/z
= 1481,6572 [M + Na]+ (calcd 1481,6565), consistent with a
molecular formula of C70H106O32Na. Its FABMS (negative-ion
mode) showed a quasi-molecular ion peak at m/z = 1457 [M
− H]−, indicating a molecular weight of 1458. Other significant
fragment ion peaks were observed at m/z = 1281 [(M − H) −176]−,
1119 [(M − H)–176 − 162]−, and 957 [(M − H)–176 − 162 − 162]−,
corresponding to the successive loss of one feruloyl moiety, and
two hexosyl moieties, respectively.
18 3.27 dd, J = 12.5, 2.5
19 1.25, 1.68 t, J = 12.5
3.35 dd, J = 12.1, 2.4
1.28, 1.70 t, J = 12.1
20
21
22
23
24
25
26
–
–
1.76, 1.95
1.69, nd
1.22 s
1.75, 1.89
1.65, nd
1.21 s
1.01 s
0.71 s
0.89 s
1.04 s
0.74 s
0.90 s
27 3.72 d, J = 10.5, 3.98
3.73 d, J = 10.0, 4.03
–
1
The H- and 13C-NMR signals of 2 assigned from the 2D NMR
28
29
30
–
spectra were almost superimposable on those of 1 except for
the acyl moiety and the position of acylation (Tables 1, 2 and 3).
Actually, inside the same oligosaccharidic chain, a substitution is
observed at GlcIV-6 position with signals at δC 63.9 and δH 4.91,
5.18, instead of GlcV-6 position in 1. Moreover, characteristic NMR
signals of a (E) feruloyl unit were found corresponding to previous
data from the literature (Table 3).[13]
0.81 s
0.93 s
0.83 s
0.96 s
Overlapped proton NMR signals are reported without designated
multiplicity.
Nd, not determined.
On the basis of the above results, the structure of com-
pound 2 was elucidated as 3-O-{β-D-glucopyranosyl-(1 → 2)-6-
O-[(E)-feruloyl]-β-D-glucopyranosyl-(1 → 3)-β-D-glucopyranosyl-
(1 → 4)-[β-D-glucopyranosyl-(1 → 2)]-β-D-glucopyranosyl}-3,27-
dihydroxyoleanolic acid (2), a new saponin named tetraptero-
side B.
(Table 1) using the correlations observed in COSY, NOESY, HSQC,
andHMBCspectra,andwasinfullagreementwithliteraturedata.[9]
For the oligosaccharidic chain, the 1H-NMR spectrum showed five
anomeric protons at δH 4.72 (d, J = 7.1 Hz), 4.91 (d, J = 7.6 Hz), 4.95
(d, J = 7.6 Hz), 5.17 (d, J = 7.8 Hz), and 5.44 (d, J = 7.6 Hz), which
gave correlations, in the HSQC spectrum, with anomeric carbon
signals at δC 104.3, 103.2, 104.3, 105.7, and 102.2, respectively.
The ring protons of the monosaccharide residues were assigned
starting from the readily identifiable anomeric protons by means
of COSY, TOCSY, NOESY, HSQC, and HMBC experiments (Table 2).
Unitsoffiveß-D-glucopyranosyl(Glc)wereidentified.Therelatively
large 3JH−1,H−2 values of the anomeric protons of Glc (7.1–7.8 Hz)
indicated a ß-anomeric orientation. The D configuration of Glc was
determined by gas chromatography (GC) analysis.[10] Correlations
observed in the HMBC spectrum between signals at δH 4.72
(GlcI-1) and δC 89.5 (C-3) confirmed the substitution at the C-3
position of the aglycone by a ß-D-glucopyranosyl moiety (GlcI).
Signals at δC 78.8 (GlcI-2) and δC 80.2 (GlcI-4) in comparison with a
terminal ß-D-glucopyranosyl moiety suggested a 2,4 substitution
of GlcI by GlcII and GlcIII, respectively. This is confirmed by cross
Experimental
General
Optical rotations values were recorded on a AA-OR automatic
polarimeter. HR-ESIMS (positive-ion mode) was carried out on a
Q-TOF 1-micromass spectrometer. FABMS were conducted in the
negative-ion mode on a Jeol SX-102 instrument. Medium-pressure
liquid chromatography (MPLC) was performed on a Gilson pump
M 305, with Bu¨chi glass column (460 mm × 25 mm and 460 mm
× 15 mm), a Bu¨chi precolumn (110 mm × 15 mm), using silicagel
60 (Merck, 15 −40 µm). Vacuum liquid chromatography (VLC)
was carried out using reversed-phase RP-18 (25 −40 µm) and
silica gel 60 (63 −200 µm) (Merck). TLC and HPTLC employed
c
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O. P. Note´ et al.
Table 2. 1H-NMR (600 MHz) and 13C-NMR (150 MHz) data of the sugar moieties of 1 and 2 in pyridine-d5
(δ in ppm, J in Hz)
Compound1
Compound 2
No
1H-NMR
13C-NMR
1H-NMR
13C-NMR
GlcI-1
4.72 d, J = 7.1
4.27 t, J = 8.3
4.23 t, J = 9.3
4.07 d, J = 9.3
3.80
104.3
78.8
77.1
80.2
75.6
62.4
102.2
74.5
77.4
71.0
77.2
61.5
104.3
74.5
90.1
70.0
78.0
61.9
103.2
84.5
76.9
70.8
78.0
62.0
4.71 d, J = 7.6
104.1
79.0
77.2
79.9
75.5
62.3
102.3
74.0
77.2
69.8
77.0
61.2
104.1
74.3
89.9
70.5
78.0
61.8
103.0
84.6
76.8
70.5
75.2
63.9
2
4.31
4.20
3
4
4.11
5
3.68
6
4.04, 4.33
5.44 d, J = 7.6
3.93
4.06, 4.38
5.50 d, J = 7.4
4.04
GlcII-1
2
3
4.15 t, J = 9.0
4.03
4.20
4
3.85 t, J = 8.1
3.81
5
3.74
6
4.32, 4.40 dd, J = 11.3, 2.0
4.95 d, J = 7.6
3.94
4.33, 4.43
5.00 d, J = 7.8
4.00
GlcIII-1
2
3
3.74
3.84 t, J = 7.4
4.08
4
3.79
5
3.90
3.95
6
4.13, 4.33
4.91 d, J = 7.6
3.96
4.20, 4.40
5.02 d, J = 7.1
4.03
GlcIV-1
2
3
4
5
6
4.22 d, J = 9.3
3.98
4.27
3.98
3.90
4.10
4.11, 4.44 br d, J = 10.2
4.91 dd, J = 11.4, 5.9
5.18 dd, J = 11.2, 6.4
5.26 d, J = 7.8
4.10
GlcV-1
5.17 d, J = 7.8
105.7
75.9
77.3
70.2
75.9
63.9
105.8
75.2
77.0
70.5
nd
2
3
4
5
6
4.03
4.17
4.25
3.79
3.99
4.04
nd
4.71, 5.13 br d, J = 11.9
4.15, 4.43
61.8
Overlapped proton NMR signals are reported without designated multiplicity.
Nd, not determined.
precoated silica gel 60F254 plates (Merck). The following TLC
solvent system CHCl3 –MeOH−AcOH −H2O (60 : 32 : 0.5 : 6.5) was
used. The spray reagent was Komarowsky reagent, a mixture (5 : 1)
of p-hydroxybenzaldehyde (2% in MeOH) and ethanolic H2SO4
(50%).
shifts are given on the δ scale and referenced to the residual
solvent signals (δH = 7.19, δC = 123.5). Coupling constants (J)
are in Hz. For 2D experiments, Varian software using pulse field
gradient were applied. The pulse conditions in C5D5N were as
follows: for the 1H-NMR spectrum, observation frequency (OF)
= 599.88 MHz, acquisition time (AQ) = 4.202 s, relaxation delay
(RD) = 5.0 s, 90 pulse width = 10.0 µs, spectral width (SW) =
7798.8 Hz, Fourier transform (FT) size = 65 536; for the 13C-
NMR spectrum, OF = 150.854 MHz, AQ = 0.453 s, RD = 1.547 s,
90 pulse width = 15.8 µs, SW = 36 182.7 Hz, line broadening
(LB) = 1.0 Hz, FT size = 65 536; for the COSY spectrum, AQ =
0.131, F2 = 2048, F1 = 256, RD = 0.369, SW = 7798.8 Hz; for
the NOESY spectrum, AQ = 0.131, F2 = 2048, F1 = 256, RD =
0.369, SW = 7798.8 Hz, mixing time = 500 ms; for the TOCSY
spectrum, AQ = 0.131, F2 = 2048, F1 = 256, RD = 0.369, SW =
7798.8 Hz, mixing time = 60 ms; for the HSQC spectrum, AQ =
0.131, RD =0.369, F1 = 36 182.7 Hz, F1 = 7798.8 Hz; for the HMBC
spectrum, AQ = 0.131, spectra frequency (SF) = 599.880 MHz,
Plant material
The stem bark of T. tetraptera was collected at Eloundem, near
Yaounde´, Cameroon, in November 2006, and identified by Dr P.
Nana, botanist of the National Herbarium of Cameroon (NHC),
Yaounde´, where a voucher specimen (No. 5567) was deposited.
NMR spectroscopy
The NMR spectra were recorded on a Varian UNITY Inova
600 spectrometer equipped with 5-mm probes. Samples were
dissolved in 160 ml of C5D5N and 20 ml of D2O and transferred
into 5-mm NMR tubes (Shigemi). The 1H and 13C-NMR spectra (at
600 and 150 MHz respectively) were measured at 303 K. Chemical
RD = 0.369, delay time (DE) = 50 ms, F1 = 36 182.7 Hz, F1
=
7798.8 Hz.
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