Lignan glucosides from sinomenium acutum rhizomes

Article abstract of DOI:10.1271/bbb.130445

The new lignan glucoside, acutumoside (1), was isolated from Sinomenium acutum rhizomes together with nine known compounds (2-10). The structure of 1 was elucidated on the basis of extensive spectroscopic analyses, including two-dimensional nuclear magnet

Full text of DOI:10.1271/bbb.130445

Biosci. Biotechnol. Biochem., 77 (10), 2144–2147, 2013
Note
Lignan Glucosides from Sinomenium acutum Rhizomes
1
1
1
1
Ki Hyun KIM, Sae Rom MOON, Chung Sub KIM, Kyeong Wan WOO,
2
1;y
Sang Un CHOI, and Kang Ro LEE
1
Natural Products Laboratory, School of Pharmacy, Sungkyunkwan University, Suwon 440-746, Korea
Korea Research Institute of Chemical Technology, Teajeon 305-600, Korea
2
Received June 3, 2013; Accepted July 18, 2013; Online Publication, October 7, 2013
[doi:10.1271/bbb.130445]
The new lignan glucoside, acutumoside (1), was
fraction (20 g) was subjected to silica gel (500 g) column
chromatography, eluting with n-hexane-ethyl acetate
(EtOAc; 10/1 to 1/1, v/v), to yield 11 fractions
(H1–H11) based on thin-layer chromatographic (TLC)
analyses. Fraction H2 (2.7 g) was subjected to octadecyl
silane (ODS, 100 g) column chromatography (MeOH–
H2O, 19:1) to give seven subfractions (H21–H27), of
which H25 was further purified by semi-preparative
high-performance liquid chromatography (HPLC), using
an Alltech Apollo silica column (250 mm ꢁ 10 mm i.d.,
isolated from Sinomenium acutum rhizomes together
with nine known compounds (2–10). The structure of 1
was elucidated on the basis of extensive spectroscopic
analyses, including two-dimensional nuclear magnetic
resonance and chemical reactions. Compounds 2, 7, 8,
and 10 displayed potential antiproliferative activity
against A549, SK-OV-3, SK-MEL-2, and HCT-15 cell
lines, while compound 1 showed weak activity against
these human tumor cells.
5
mm; n-hexane-EtOAc ¼ 18:1, v/v at a flow rate of
Key words: Sinomenium acutum; Menispermaceae;
lignan; acutumoside; cytotoxicity
2 mL/min) equipped with a Shodex refractive index
detector, to afford 7 (25 mg). Fraction H3 (5.0 g) was
successively subjected to Sephadex LH-20 column
chromatography (CH Cl –MeOH, 1:1, v/v) and silica
Sinomenium acutum (Thumb.) Rehd. et Wils.
Menispermaceae) is a deciduous shrub distributed in
2
2
(
gel (100 g) column chromatography to obtain four
subfractions (H31–H34), of which H32 was further
purified by semi-preparative HPLC, using an Alltech
Econosil RP-18 column (250 mm ꢁ 10 mm i.d., 10 mm;
100% MeOH at a flow rate of 2 mL/min) equipped with
a Shodex refractive index detector, to yield compounds
8 (6 mg) and 9 (5 mg). Compound 10 (26 mg) was
isolated from fraction H4 (1.7 g) by passage through a
Sephadex LH-20 column (CH Cl –MeOH, 1:1, v/v) and
Asia, particularly in Japan, China, and Korea. The
rhizome of S. acutum, known as qing-feng-teng, is a
Chinese medicine commonly used for the treatment of
1
)
rheumatism, neuralgia, arthritis, and edema. The
characteristic constituents of this plant are such alka-
loids as morphinane, hasubanane, bisbenzylisoquinoline,
2
–6)
aporphine, and protoberberine alkaloids.
main component, sinomenine, displays marked biolog-
Of these, the
2
2
7
)
ical activities including arthritis amelioration, immu-
9)
separation using the same HPLC system (n-hexane-
EtOAc ¼ 6:1). The n-BuOH-soluble fraction (20 g) was
subjected to silica gel (500 g) column chromatography,
eluting with CHCl3-MeOH (20:1–1:1, v/v), to yield
eight fractions (B1–B8) based on TLC analyses.
Fraction B5 (2.4 g) was subjected to silica gel (100 g)
column chromatography (CHCl3–MeOH, 10:1) and then
to ODS (100 g) column chromatography (MeOH–H2O,
2:3) to give four subfractions (B51–B54), of which B53
was further purified by semi-preparative HPLC (MeOH–
8)
nomodulation, and vasodilation.
In our continuing search for new bioactive substances
from Korean medicinal plants, we investigated the
chemical constituents of the rhizomes of S. acutum.
We isolated a new lignan glucoside, named acutumoside
1), together with nine known compounds (2–10)
Fig. 1A) from the rhizomes. Their structures were
elucidated on the basis of extensive spectroscopic
analyses, including two-dimensional nuclear magnetic
resonance (2D NMR) and chemical reactions. This
report describes the isolation and structural elucidation
of new compound 1, as well as the antiproliferative
activities of the isolates (1–10) toward human cancer
cell lines.
(
(
H O, 2:3) to yield compounds 1 (26 mg), 2 (15 mg), and
2
3 (66 mg). The purification of fraction B6 (5.0 g) by
repeated column chromatography (7:1 CHCl –MeOH,
3
and 2:3 MeOH–H O) and HPLC separation (3:7
2
MeOH–H2O) resulted in the isolation of compounds 4
(13 mg), 5 (5 mg), and 6 (19 mg).
Compound 1 was isolated as a colorless gum with
The dried rhizomes of S. acutum (3.5 kg) were
chopped and extracted with 80% aqueous methanol
ꢀ
25
D
(
MeOH) three times daily at 60 C and filtered. The
negative optical rotation (½ꢀꢂ ꢃ83.1, MeOH). The
filtrate was then concentrated under vacuum to afford a
crude MeOH extract (390 g) which was suspended in
H2O and successively partitioned with n-hexane, chloro-
form (CHCl ), and n-butanol (BuOH) to respectively
3
yield 21, 21, and 65 g of a residue. The n-hexane-soluble
molecular formula of 1 was determined as C25H32O10
from its positive mode high-resolution electron spray
ionization mass spectroscopic data at m=z 515.1895
þ
½M þ Naꢂ (calcd. for C H NaO , 515.1893), which
25 32
10
was compatible with the nuclear magnetic resonance
y
To whom correspondence should be addressed. Tel: +82-31-290-7710; Fax: +82-31-290-7730; E-mail: krlee@skku.ac.kr
Abbreviations: SRB, sulforhodamine B

Lignans from Sinomenium acutum
2145
5
2
A
HO
OH
6
9
H CO
4
HO
HO
3
6
'
7'
9'
OH
7
5'
3
1
8
H CO
1'
2'
OH
3
8'
4
'
H3CO
O
4
''
O
1
5
''
O
3'
OH
OCH3
OCH3
''
HO
HO
2
''
OCH3
OH
3
''
2
1
OCH3
OH
OR2
O
O
OCH3
OCH3
OH
OH
HO
HO
H CO
H3CO
R1O
3
O
O
HO
OCH3
3
OCH3
4: R1 = H, R2 = Glc
5: R1 = Glc, R2 = Glc
HO
OH
GlcO
O
H CO
3
OCH3
6
O
7
HO
HO
HO
8
9
10
HO
H CO
OH
B
OH
3
O
HO
HO
O
OH
OCH3
Fig. 1. Structures of Compounds 1–10.
A, Structures of compounds 1–10. B, Selected H- H COSY (bold lines) and HMBC (arrow) correlations for 1.
1
1
0
(
NMR) data. The IR spectrum of 1 showed the presence
ꢃ1
group was located at the C-4 position of the aglycone
skeleton (Fig. 1B). The gross structure of 1 was
of hydroxyl (3420 cm ) and aromatic groups (1615 and
450 cm ). The 1H- and
Table 1) of 1 resembled those of icariside E5, with
the exception of resonances due to a sugar moiety [ꢁH
.98 (1H, d, J ¼ 8:0), 3.84 (1H, m), 3.82 (1H, m), 3.32
1H, m), 3.22 (1H, dd, J ¼ 8:5, 7.5), and 3.17 (1H, dd,
J ¼ 11:0, 10.5); ꢁC 103.3, 76.5, 73.5, 69.8, and 65.5] in
. Acid hydrolysis of 1 was conducted by heating a
ꢃ1
13
C-NMR spectral data
determined by H- H correlation spectroscopy (COSY),
heteronuclear multiple-quantum coherence (HMQC),
and HMBC experiments, the key H- H COSY and
HMBC correlations being given in Fig. 1B. The abso-
lute configuration of C-8 was established by an analysis
of the optical rotation of aglycone 1a obtained from the
1
1
1
(
10)
1
1
4
(
2
5
1
acid hydrolysis. The optical rotation value (½ꢀꢂ ꢃ15.8,
D
solution of 1 in 1 N HCl (dioxane/H O ¼ 1:1, 5 mL)
MeOH) of 1a suggested it to have the 8R configuration
by a comparison of the optical rotation of 1a with that
2
under reflux for 2 h, affording aglycone 1a and a sugar
moiety.1 Aglycone 1a was extracted with EtOAc and
confirmed as isodehydrodiconiferyl alcohol on the basis
1)
25
D
pound, (8R)-isodehydrodiconiferyl alcohol, in the liter-
(½ꢀꢂ ꢃ18.4, MeOH) of the previously reported com-
1
10,12)
of an analysis of its MS and H-NMR data (Table 1) and
ature. On the basis of the foregoing data, the
structure of 1 was determined to be (8R)-isodehydrodi-
coniferyl alcohol-4 -ꢂ-D-xylopyranoside and named
1
0)
by a comparison with reported spectroscopic data. The
sugar moiety obtained from the aqueous layer was
analyzed as D-xylose by silica gel co-TLC in comparison
with an authentic sample [CHCl3/MeOH/H2O solvent
system (8:5:1); TLC (Rf 0.56)] and by comparing the
0
1
3)
acutumoside.
The known compounds were identified as (1R)-
(4-hydroxy-3-methoxyphenyl)-(2R)-[4-[(1E)-3-hydroxy-
1-propen-1-yl]-2,6-dimethoxyphenoxy]-1,3-propanediol
2
D
5
sign of its specific rotation value (½ꢀꢂ þ20.5, H O).
2
1
4)
15)
The heteronuclear multiple-bond correlation (HMBC)
spectrum showed that the signal for an anomeric proton
at ꢁH 4.98 had long-range correlation with the carbon
(2), (ꢃ)-syringaresinol (3), (þ)-syringaresinol-O-ꢂ-
16,17) 18)
D-glucopyranoside (4),
liriodendrin (5), (7S,8R)-
dehydrodiconiferyl alcohol 4-O-ꢂ-D-glucopyranoside
(6), lupenone (7), 3-epi-lupeol (8), 3ꢂ-hydroxy-
0
19)
20)
21)
resonance at ꢁC 143.5 (C-4 ), suggesting that the xylosyl

2146
K. H. KIM et al.
Table 1. 1H-(500 MHz) and 13C-(125 MHz) NMR Data for Com-
pounds 1 and 1a in CD3OD (ꢁ in ppm)
Table 2. Antiproliferative Activities of Compounds 1–10 against
Four Cultured Human Tumor Cell Lines
a
1
1a
IC50 (mM)
No.
Compound
ꢁ
H
ꢁC
ꢁH
A549
SK-OV-3
SK-MEL-2
HCT-15
1
2
3
4
5
6
7
132.3
113.0
147.3
144.3
114.6
121.6
37.0
1
25.87
10.03
>30.0
>30.0
>30.0
>30.0
>30.0
12.16
>30.0
>30.0
0.01
29.10
8.41
26.53
7.38
>30.0
18.83
>30.0
>30.0
>30.0
>30.0
>30.0
11.93
>30.0
>30.0
0.13
6.74 br s
6.57 d (1.5)
2
3
>30.0
>30.0
>30.0
>30.0
>30.0
11.84
>30.0
21.55
0.02
>30.0
>30.0
>30.0
>30.0
18.63
9.73
4
6.65 br s
6.65 br s
6.56 d (8.0)
5
6.47 dd (8.0, 1.5)
2.95 dd (13.5, 6.0);
2.70 dd (13.5, 9.0)
3.95 m
6
2.93 dd (13.5, 6.5);
.85 dd (13.5, 9.0)
3.81 m
7
2
8
8
9
1
2
3
4
5
6
7
8
9
3
3
1
2
3
4
5
42.3
64.7
9
10
>30.0
13.14
0.01
3.78 m; 3.62 m
3.75 m; 3.68 m
0
0
0
0
0
0
0
0
0
b
133.6
108.5
152.0
143.5
137.4
119.0
130.3
128.2
62.5
Doxorubicin
6.94 d (2.0)
6.91 d (2.0)
aIC50 value of compounds against each cancer cell line, which was defined
as the concentration (mM) that caused 50% inhibition of cell growth in vitro.
Doxorubicin as a positive control.
b
6.89 d (2.0)
6.54 dd (15.5, 1.0)
6.27 dt (15.5, 6.0)
4.21 dd (6.0, 1.0)
3.76 s
6.90 d (2.0)
6.53 dd (15.5, 1.0)
6.29 dt (15.5, 5.5)
4.21 dd (5.5, 1.0)
3.71 s
investigate non-alkaloidal compounds from S. acutum
suppressing the survival of cancer cells.
-OCH3
0
55.4
55.3
Acknowledgments
-OCH3
3.85 s
3.85 s
0
0
0
0
0
0
0
0
0
0
4.98 d (8.0)
3.22 dd (8.5, 7.5)
3.32 m
103.3
73.5
76.5
This work was supported by grant no. 09112KFDA890
from Korea Food and Drug Administration in Korea.
We thank Drs. E. J. Bang, S. G. Kim, and J. J. Seo at
the Korea Basic Science Institute for their assistance
with the NMR spectroscopic and mass spectrometric
measurements.
3.84 m
3.82 m;
69.8
65.5
3.17 dd (11.0, 10.5)
ꢄ
Assignments were based on 1H- H COSY, HMQC, and HMBC experi-
1
ments. Well-resolved couplings are expressed with coupling patterns and
coupling constants in Hz in parentheses.
References and Notes
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N HCl (dioxane/H2O, 1:1, 5 mL) under the reflux condition
1
1
and SK-MEL-2 cells, with IC50 values ranging from
for 2 h. After cooling, the reaction mixture was diluted with
H O and then extracted with an equal amount of EtOAc (ꢁ3),
2
5.87 to 29.10 mM.
In conclusion, we isolated and identified the new
2
the EtOAc layer being evaporated under reduced pressure. The
resulting residue was dried and purified in a Waters silica gel
Sep-Pak Vac 6 cc column (CHCl3–MeOH, 15:1) to give
aglycone 1a (3.5 mg). This aglycone was identified as (8R)-
isodehydrodiconiferyl alcohol by comparing its 1H-NMR, MS,
and specific rotation with the data reported in the literature.
The aqueous fraction was neutralized by passage through an
Amberlite IRA-67 column and was repeatedly evaporated under
lignan glucoside, acutumoside (1), together with nine
known compounds (2–10) from S. acutum rhizomes and
evaluated their antiproliferative activities against four
human tumor cell lines. Alkaloids have hitherto been
studied with keen interest as the antitumor constituents
of S. acutum. To our knowledge, this is the first report to

Lignans from Sinomenium acutum
2147
reduced pressure to give a sugar fraction. This sugar fraction
was subjected to column chromatography in a silica gel column,
eluting with MeCN–H2O (8:1) to yield D-xylose with a positive
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25
specific rotation (½ꢀꢂ þ20.5, c 1.5 in H2O). This D-xylose was
D
analyzed by silica gel co-TLC in comparison with an authentic
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D
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25
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1
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1
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