G Model
PHYTOL 552 1–6
O.P. Note´ et al. / Phytochemistry Letters xxx (2013) xxx–xxx
3
Table 1
59
60
61
which indicate of the loss of two pentosyl units. Extensive analysis
of 1D and 2D NMR spectra (1H NMR, 13C NMR, DEPT, COSY, NOESY,
HSQC and HMBC) indicated the presence of seven tertiary methyl
NMR spectroscopic data (600 MHz for 1H and 150 MHz for 13C) for the aglycone
moieties of compounds 1and 2 (
d
in ppm and J in Hz)a.
1
2
62
63
64
65
66
67
68
69
70
71
72
73
74
75
76
77
78
79
80
81
82
83
84
85
groups at
broad triplet proton at
carbon at 122.5 (C-12), a quaternary carbon at
oxymethine protons at 3.26 (brd, J = 7.6 Hz, H-3) which are
typical signals of an olean-12-ene skeleton. The aglycone moiety of
1 was thus recognized to be oleanolic acid by 1H NMR and 13C NMR
analyses (Table 1) using the correlations observed in COSY, NOESY,
HSQC, and HMBC spectra, and was in full agreement with literature
data (Carpani et al., 1989; Nigam et al., 1997; Mimaki et al., 2004).
d
0.70, 0.82, 0.83, 0.86, 0.87, 1.18, and 1.19, an olefinic
5.31 (brt, J = 3.5 Hz, H-12) coupled to a
143.8 (C-13), one
d
No C
dC
dH
dC
dH
d
d
1
38.4
1.39b
46.4
1.05; 2.12 (d, 4.0)
d
2
25.9
89.2
38.8
55.5
18.0
32.8
38.8
47.6
36.5
23.1
122.5
143.8
42.0
28.2
23.1
46.7
41.6
45.8
30.3
32.8
31.6
27.8
16.6
15.0
16.6
25.9
176.4
32.8
23.2
1.64; 2.03
3.26 (brd, 7.6)
–
66.8
94.6
40.5
55.0
18.3
32.8
39.4
47.6
37.3
23.8
123.0
143.2
41.9
36.0
27.5
47.1
41.7
45.6
30.5
33.3
32.8
27.9
17.8
15.2
17.5
25.6
176.3
32.7
23.1
3.62 (ddd, 11.2, 9.2, 4.0)
3
3.20 (dd, 9.2, 11.2)
4
–
5
0.64
0.64
6
1.22; 1.46
1.39b
1.12; 1.36
7
1.36; 1.46
8
–
–
9
1.53
1.56
The chemical shifts of C-3 (d 89.2) and C-28 (d 176.4) (Table 1)
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
–
–
indicated that 1 is a bidesmosidic glycoside (Woldemichael and
Wink, 2001; Sahu and Achari, 2001) of oleanolic acid with sugar
chains linked to C-3 and C-28 of the aglycone through an ether and
ester bond, respectively.
The 1H NMR spectrum of compound 1 showed eight anomeric
signals at d 4.97 (d, J = 7.1 Hz), 4.92 (d, J = 7.6 Hz), 4.99 (d,
J = 2.9 Hz), 4.83 (d, J = 7.6 Hz), 5.96 (d, J = 8.1 Hz), 5.28 (d,
J = 7.6 Hz), 6.12 (d, J = 2.1 Hz), and 5.91 (d, J = 3.8 Hz), which
correlated with eight anomeric carbon atom resonances at d 104.0,
104.8, 102.6, 105.9, 93.0, 104.3, 110.8, and 110.2, respectively in
the HSQC spectrum (Table 2). From the anomeric proton of each
monosaccharide moiety, all the protons within each spin system
were assigned by means of COSY, NOESY, HSQC, and HMBC
1.83b
1.84
5.31 (brt, 3.5)
–
5.34 (brt, 3.2)
–
–
–
1.11; 2.06
1.79
1.03; 1.83
0.82
–
–
3.03 (d, 11.6)
1.13; 1.67
–
1.39b
3.06
1.12; 1.66
–
b
1.57; 1.81
1.18 (s)
0.87 (s)
0.70 (s)
0.86 (s)
1.19 (s)
–
1.36; 1.46
1.22 (s)
0.81 (s)
0.77 (s)
0.79 (s)
1.12 (s)
–
experiments. Units of one 2-acetamido-2-deoxy-
(GlcNAc), one -glucopyranosyl (Glc), two -xylopyranosyl (Xyl I
and Xyl II), two -apiofuranosyl (Api I and Api II), and one - and
b-glucopyranosyl
86
87
88
b
b
b
a
b-
0.83 (s)
0.82 (s)
0.86 (s)
0.82 (s)
arabinopyranosyl (Ara I and Ara II, respectively), were identified
89
90
91
92
(Table 2). The anomeric protons of Ara I were determined to have
Assignments were based on the HMBC, HSQC, COSY and DEPT experiments.
aOverlapped 1H NMR signals are reported without designated multiplicity.
bnot determined.
3
the
a
-orientation based on its relatively large JH-1
,
value of
H-2
7.6 Hz, whereas Ara II was
b
-orientated based on its relatively
3
small JH-1,
H-2
value of 2.9 Hz (Tene et al., 2011). The absolute
93
configuration of these sugar moieties were determined to be
D for
94
GlcNAc, Api, and Xyl, and for Ara by GC analysis (Section 3). The
L
95
96
97
98
sequencing of the glycoside chains were achieved by analysis of
HMBC and NOESY experiments. The cross peak correlations
NOESY correlation observed between H-3 (
d
4.04) of Xyl II and H-1 125
6.12) of Api I, and the HMBC correlation observed between H-1 126
H 5.91) of Api II and C-4 ( C 79.5) of Glc, were useful to attach Api 127
(
(
d
d
observed in the HMBC spectrum between H-1 (
and C-3 ( 89.2) of the aglycone, and in the NOESY spectrum
between H-1 ( 4.97) of GlcNAc and H-3 ( 3.26) of oleanolic acid,
suggested that GlcNAc was directly attached to C-3 of the aglycone.
Moreover, the HMBC correlation observed between H-1 ( 4.92) of
Ara I and C-3 ( 79.5) of GlcNAc established the connectivity
between the two sugar units, which was confirmed by the reverse
HMBC correlation observed between H-3 ( 4.34) of GlcNAc and C-
1 ( 104.8) of Ara I. On the other hand, the HMBC correlation
observed between H-1 ( 4.99) of Ara II and C-6 ( 67.8) of GlcNAc
allowed us to locate Ara II at C-6 of GlcNHAc. This was supported by
the NOESY correlation observed between H-1 ( 4.99) of Ara II and
H-6a ( 4.10) of GlcNAc. This Ara II was substituted at its C-2 by Xyl
I, as evidenced by the direct and reverse correlations observed in
the HMBC spectrum between H-2 ( 4.38) of Ara II and C-1 ( 105.9)
of Xyl I, and between H-1 ( 4.83) of Xyl I and C-2 ( 80.5) of Ara II
d
4.97) of GlcNAc
d
d
I and Api II at C-3 and C-4 of Xyl II and Glc, respectively. In the same 128
way, the absence of any 13C NMR glycosylation shifts for Api I and 129
Api II supported their terminal positions. Thus, the tetrasacchar- 130
99
d
d
100
101
102
103
104
105
106
107
108
109
110
111
112
113
114
115
116
117
118
119
120
121
122
123
124
d
ide
apiofuranosyl -(1!4)]-
lished to be linked at C-28 of the aglycone (Fig . 2). Based on 133
b
-
D
-apiofuranosyl-(1!3)-
b
-
D
-xylopyranosyl-(1!2)-[
b
-
D
-
131
d
b-D
-glucopyranosyl moiety was estab- 132
d
the above evidences, the structure of 1 was established as 3-O-
-xylopyranosyl- (1!2)- -arabinopyranosyl-(1!6)-[
b
a- L
-
-
134
135
d
[
b
-
D
a
-
L
d
d
arabinopyranosyl-(1!3)]-2-acetamido-2-deoxy-
b
-
D
-glucopyra- 136
nosyl]-
28-O-[
-a1piofuranosyl -(1!4)]-
b
-
D
-apiofuranosyl-(1!3)-
-glucopyranosyl]-olea-
b- D-xylopyranosyl- 137
d
(1!2)-[
b
-
D
b
-
D
138
139
d
nolic acid.
By extensive analysis of their NMR data (1H, 13C NMR, DEPT, 140
COSY, NOESY,HSQCandHMBC)andmassspectrometry,compounds 141
d
d
d
d
2 and 3 were identified as 3-O-
arabinopyranosyl-(1!3)- -glucopyranosyl]maslinic
-glucopyranosyl-(1!2)- -rhamnopyranosyl] ester and 3-O- 144
-arabinopyranosyl-(1!2)-
glucopyranosyl]maslinic acid-28-[
b-[
a
-
L
-arabinopyranosyl-(1!2)-
a
-
L
-
142
(Fig. 2). The terminal positions of Xyl I and Ara I was evidenced by
the absence of any 13C NMR glycosylation shifts for these sugar
b-D
acid-28- 143
[b-D
a-L
moieties. Thus, the tetrasaccharide
arabinopyranosyl-(1!6)-[ -arabinopyranosyl-(1!3)]-2-aceta-
mido-2-deoxy- -glucopyranosyl moiety was established to be
linked at C-3 of the aglycone (Fig. 2). Furthermore, the cross peak
observed in the HSQC spectrum at 5.96/ 93.0 (Glc H-1/C-1)
b-
D-xylopyranosyl-(1!2)-
a
-L-
b
-[
a
-
L
a
-
b
L
-arabinopyranosyl-(1!3)-
b
b-D-
-
D
-
145
146
a
-L
-D
-glucopyranosyl-(1!6)-
b
-
D
glucopyranosyl-(1!2)-
a-L
-rhamnopyranosyl] ester, respectively 147
(Tchivounda et al., 1991).
148
d
d
In conclusion, the present study is the first report on the 149
saponin content of P. africanum, and the presence of the two known 150
compounds (2 and 3) in this species, previously isolated from 151
Cylicodiscus gabunensis (Tchivounda et al., 1991) may indicate a 152
close relationship between the two species of Mimosaceae 153
suggested that this sugar should be directly attached to C-28
through an ester bond. The correlations observed in the HMBC
spectrum between H-2 (
and in the NOESY spectrum between H-2 (
5.28) of Xyl II allowed us to locate Xyl II at C-2 of Glc. Moreover, the
d
4.20) of Glc and C-1 (
d 104.3) of Xyl II,
d
4.20) of Glc and H-1 (
d
subfamily.
154
´
Please cite this article in press as: Note, O.P., et al., Triterpenoid saponins from Piptadeniastrum africanum (Hook. f.) Brenan. Phytochem.