M. Thapsut et al.
Phytochemistry Letters 44 (2021) 42–47
1
1
1
1
tertiary carbons and one keto carbon. The H- H COSY spectrum showed
correlations of H-7/H-8/H-9/H-10. The 3-oxo-megastigman-7-ene
moiety was established from the HMBC spectrum which exhibited
cross-peaks between H-2/C-1, C-3, C-4, C-6, C-11, C-12; H-4/C-2, C-3, C-
MeOH). The successive H- H connectivities from H-2, H-3 and H-4 and
H-7, H-8, H-9 and H-10 were revealed from its 1H, H COSY spectrum.
The trans-double bond between C-7/C-8 was proposed from the JH-7/H-8
value of 15.9 Hz. The presence of a β-glucopyranosyl moiety was evident
in particular from the 1H and C NMR chemical shifts of an anomeric
1
13
5
, C-6, C-13; H-7/C-1, C-5, C-6, C-8, C-9; and H-8/C-6, C-7, C-9, and C-10
1
13
(
Fig. 2). The presence of a tetrahydrofuran moiety with the ether linkage
group at δ
H
4.41 (d, J =8.0 Hz) and δ
C
101.8. Most of the H and
C
between C-5 and C-11 was revealed from the HMBC cross-peaks between
H-11/C-1, C-2, C-5, C-6, and C-12. The coupling constant JH-7/H-8 with a
value of 15.9 Hz indicated a trans-double bond between C-7/C-8. Based
on the above data, compound 1 was proposed to have a planar structure
similar to that of drummondol (Powell and Smith, 1981). The absolute
configuration, particularly at C-9, of drummondol first obtained from
NMR resonances were very similar to those of (3S,5R,6R,7E,9S)-mega-
stigman-7-ene-3,5,6,9-tetrol-3-O-β-D-glucopyranoside, which was also
isolated in this study. However, H-3 was found to resonate at δ
H
4.17 as
quintet-like (J =3.5 Hz) instead of at ca. δ
H
4.19 as triplet of triplets (J =
11.7 and 4.3 Hz) as previously reported (Otsuka et al., 2003) and found
for (3S,5R,6R,7E,9S)-megastigman-7-ene-3,5,6,9-tetrol-3-O-β-D-gluco-
pyranoside. The NOESY spectrum showed similar NOE correlations as
those found in (3S,5R,6R,7E,9S)-megastigman-7-ene-3,5,6,9-tetro-
l-3-O-β-D-glucopyranoside except that the NOE cross-peak between
2
3
Sesbania drummondii, [
α]
D
ꢀ 21 (c 1.03, MeOH), was not established
(
Powell et al., 1986; Powell and Smith, 1981). Later, drummondol ob-
2
5
tained from Cynanchum liukiuense, having [
α
]
D
+2.7 (c 0.30, MeOH),
was reported as 1R,5R,6S,7E,9R-drummondol (Abe et al., 2000), while
related compound (spionoside B,
3
H-3/H -12 was missing (Fig. 3). The above evidence indicated that these
the β-D-glucopyranoside of
a
two compounds possess different configurations at C-3. Direct acid hy-
drolysis of 3 was not performed due to the scarcity of 3. However, on the
basis of the result obtained from acid hydrolysis of (3S,5R,6R,7E,
9S)-megastigman-7-ene-3,5,6,9-tetrol-3-O-β-D-glucopyranoside, which
provided D-glucose, compound 3, i.e., heterophylloside B, was thus
(
9S)-drummondol-9-O-β-D-glucopyranoside) isolated from Capparis spi-
20
nosa fruits, having [
α]
D
ꢀ 51.2 (c 2.0, MeOH), gave drummondol that
was reported to have a 1R,5R,6S,7E,9S configuration after hydrolysis
Çalı s¸ et al., 2002). Assignments of the configuration at C-9 of drum-
(
mondol from both reports were based on a modified Mosher’s ester
method. In the present study, the NOESY spectrum of compound 1
concluded
to
be
(3R,5R,6R,7E,9S)-megastigman-7-ene-3,5,6,
9-tetrol-3-O-β-D-glucopyranoside.
revealed NOE interactions between H-7/H
a
-11,
H
3
-12,
H
3
-13;
Among the nine compounds, (3S,5R,6R,7E,9S)-megastigman-7-ene-
H
β
-2/H
b
-11, H
3
-12 and H
2
-4/H
3
-13, as shown in Fig. 3. On the basis of
3,5,6,9-tetrol,
2
3
the optical rotation of 1 with [
α
]
D
+15.4 (c 0.24, MeOH) and a com-
blumenol
A
(vomifoliol), (3S,5R,6R,7E,9S)-megastigman-7-ene-
(3S,5R,6R,7E)-3,5,6-trihy-
parison of the experimental and calculated ECD spectra (Fig. 4), com-
pound 1 was thus proposed as (1S,5S,6S,7E,9S)-drummondol and
named heterophyllol.
3,5,6,9-tetrol-3-O-β-D-glucopyranoside,
droxy-7-ene-9-oxo-megastigmane, (6S)-dehydrovomifoliol, ent-kaur-
ane-3β,16β,17-triol, 7,11,15-trimethyl-3-methylene-hexadecan-1,2-diol
Compound 2 was obtained as a sticky liquid with molecular formula
(phytene-1,2-diol), stigmast-4-en-3
tained in sufficient quantity, only glut-5-en-3β-ol showed IC50 values of
M, while the positive control compound
α,6β-diol and glut-5-en-3β-ol ob-
1
3
19 30 9
C H O based on its HRESIMS. The C NMR spectrum showed 19
carbon resonances. The H and 13C NMR spectra of 2 showed similar
1
9.72 ± 0.58 and 9.86 ± 1.07
μ
patterns of resonances as those found in 1, with additional resonances of
(doxorubicin) exhibited IC50 values of 0.43 ± 0.01 and 0.16 ± 0.01
μM
a β–glucopyranosyl group showing a dioxygenated methine group at δ
H
against HelaS3 and KB cells, respectively.
4
3
.34 (d, J =7.8 Hz) and δ
C
102.8 and oxymethylene resonances at δ
H
.87, 3.63, and δ
C
62.9. The HMBC spectrum showed cross-peaks be-
3. Experimental section
′
′
tween H-9/C-1 as well as H-1 /C-9, indicating connectivity of C-9-O to
′
C-1 of the glucosyl group (Fig. 2). To specify whether the glucose is D- or
3.1. General experimental procedures
L-, acid hydrolysis of the glucoside is needed. Although direct acid hy-
drolysis of 2 could not be carried out due to the scarcity of 2, D-glucose
Melting points were measured using an Electrothermal melting point
apparatus and are uncorrected. Optical rotations were recorded on a
JASCO DIP 1020 polarimeter, and ECD spectra were recorded on a
JASCO J-810 apparatus. The IR spectra were obtained on a Perkin-Elmer
1760x FT-IR spectrophotometer. The 1H and C NMR spectra were
recorded with Bruker AVANCE 400 MHz and Bruker AVANCE III HD 400
MHz NMR spectrometers. Chemical shifts are referenced to the residual
2
5
23
[
α
]
D
+16.2 (c 0.40, H
2
O); lit. [
α
]
D
+53.2 (c 0.1, H
2
O) (Perrone et al.,
2
9
012) was obtained after (3S,5R,6R,7E,9S)-megastigman-7-ene-3,5,6,
-tetrol-3-O-β-D-glucopyranoside was hydrolysed with 5% HCl. It was
13
thus assumed that all glucose found in this study is D-glucose. Based on
2
3
the optical rotation of 2, [
α
]
D
+4.1 (c 0.91, MeOH), as compared to
2
0
spionoside B, [
α]
D
ꢀ 51.2, and the co-occurrence of 1 in this plant,
compound 2 was thus elucidated as (1S,5S,6S,7E,9S)-drummondo-
l-9-O-β-D-glucopyranoside and given the name heterophylloside A.
solvent signals (MeOH-d
4
: δ
H
C
3.30 and δ 49.0 ppm). HRESIMS was
recorded on a Bruker DaltonicsmicroTOF mass spectrometer. Column
chromatographic separations were performed using silica gel 60 (less
than 0.063 mm and 0.063ꢀ 0.200 mm, Merck) and silica gel 60 RP-18
Compound 3 was obtained as an amorphous solid. Its HRESIMS
+
indicated a molecular formula of C19
H
34
O
9
based on [M + Na] at m/z
2
3
4
29.2123 (calcd for C19 Na, 429.2095) with [
H
34
O
9
α
]
D
-14.2 (c 0.22,
(40ꢀ 63
μm, Merck), Dianion HP-20 (Supelco Analytical, Bellefonte,
Fig. 1. Structures of compounds 1-3.
4
3