Leonurosides A-D: Steroid N-acetylglucosaminides from the fruits of Leonurus japonicus

Article abstract of DOI:10.1016/j.phytol.2014.10.016

Four new naturally occurring steroidal glycosides (leonurosides A-D, 1-4) were isolated from the dried fruits of Leonurus japonicus (motherwort fruit). Their structures were established by spectroscopic and chemical methods. The monosaccharide unit was identified to be a 2-acetamido-2-deoxy-β-d-glucose and the aglycone is either a common 3β-stigmasterol or 3β-ergosterol derivative. These compounds are the first representatives of steroid N-acetylglucosaminides from vascular plants.

Full text of DOI:10.1016/j.phytol.2014.10.016

Phytochemistry Letters 10 (2014) 287–290
Contents lists available at ScienceDirect
Phytochemistry Letters
journal homepage: www.elsevier.com/locate/phytol
Leonurosides A–D: Steroid N-acetylglucosaminides from the fruits
of Leonurus japonicus
a,b,c
a
c
a
a,c,
Miao Ye
, Guang-Lei Ma , Jing-Jing Su , Juan Xiong , Jin-Feng Hu
*
a
Department of Natural Products Chemistry, School of Pharmacy, Fudan University, Shanghai 201203, PR China
School of Pharmacy, Fujian University of Traditional Chinese Medicine, Fuzhou, Fujian 350122, PR China
b
c
Department of Chemistry and Institutes for Advanced Interdisciplinary Research, East China Normal University (ECNU), Shanghai 200062, PR China
A R T I C L E I N F O
A B S T R A C T
Article history:
Four new naturally occurring steroidal glycosides (leonurosides A–D, 1–4) were isolated from the dried
fruits of Leonurus japonicus (motherwort fruit). Their structures were established by spectroscopic and
Received 29 May 2014
Received in revised form 10 October 2014
Accepted 14 October 2014
Available online 25 October 2014
chemical methods. The monosaccharide unit was identified to be a 2-acetamido-2-deoxy-
b-D-glucose
and the aglycone is either a common 3 -stigmasterol or 3 -ergosterol derivative. These compounds are
b
b
the first representatives of steroid N-acetylglucosaminides from vascular plants.
ß 2014 Phytochemical Society of Europe. Published by Elsevier B.V. All rights reserved.
Keywords:
Leonurus japonicus
Lamiaceae
Steroid N-acetylglucosaminides
Leonurosides
1
. Introduction
describe the isolation and structural elucidation of the new
compounds.
Naturally occurring steroid N-acetylglucosaminides are interest-
ing secondary metabolites that generally have a 2-acetamido-2-
deoxy- -glucose moiety bound to a steroid nucleus. They have
2. Results and discussion
b-D
been often found from the excretion of some animals [e.g., the urine
or serum of mammals (Marschall et al., 1989; Meng et al., 1996,
Compound 1 was obtained as a white, amorphous powder. Its
molecular formula was established as C37
H
63NO
6
based on a
pseudomolecular ion peak [M+Na] at m/z 640.4556 (calcd for
Na, 640.4548) in its HRESIMS. In addition to eight
characteristic signals attributed to an N-acetylglucosamine moiety
100.5, 57.8, 76.1, 72.2, 78.2, 62.5; N-Ac: 170.3, 23.2) (Iida et al.,
1
997; Yamaga and Kohara, 1994) and the defense secretion of fishes
+
(Tachibana et al., 1984, 1985)], and only a few examples were
37 6
C H63NO
previously reported from marine organisms (Gulavita et al., 1994;
Lin et al., 2010). However, none of this class of steroidal glycosides
has been so far isolated from any vascular plants. During our
continuing research toward the discovery of novel bioactive sterol-
type derivatives from terrestrial higher plants (Wu et al., 2009; Ye
et al., 2013) and fungi (Zang et al., 2013), four (1–4) new steroid N-
acetylglucosaminides (Fig. 1) were identified and characterized
from the air-dried ripe fruits of the herbaceous plant Leonurus
japonicus Houtt (family Lamiaceae). In this study, compounds 3 and
(
2
2
d
d
1
3
001), the C and DEPT NMR (Table 1) spectra of 1 exhibited
9 carbon signals. Acid hydrolysis of 1 (see Section 3) gave the
aglycone 1a and the sugar moiety (existed as glucosamine
hydrochloride). 1a was ascertained to be -sitosterol by comparing
b
its NMR data (Ahmad et al., 2010; Wright et al., 1978) and optical
rotation (Gutierrez-Lugo et al., 2004) with those published. The
aglycone was also confirmed by a direct TLC analysis with an
authentic specimen. Meanwhile, the HMBC correlations between
4
were obtained as a mixture of C-24 epimers of 3-
b-O-(2-
acetamido-2-deoxy- -glucopyranosyl)-ergosta-5-ene. We herein
b-D
the acetyl carbonyl (
d 170.3) and d 8.91 (NH)/d 2.15 (3H, s)
indicated the presence of an acetamido group in the sugar unit.
Furthermore, the 2-acetamido-2-deoxysugar moiety was con-
firmed by the HMBC correlation from NH (
d
8.91, D
2
O exchange-
*
Corresponding author at: Department of Natural Products Chemistry, School of
0
able) to C-2 (
d
57.8) and the COSY correlations between NH (
d
Pharmacy, Fudan University, Shanghai 201203, PR China. Tel.: +86 21 51980172;
fax: +86 21 51980172.
0
8.91) and H-2 (
d
4.50), especially the long-range ‘‘W’’ coupling
0
E-mail address: jfhu@fudan.edu.cn (J.-F. Hu).
between NH (d 8.91) and H-1 (d 5.28) (see Supplementary data).
http://dx.doi.org/10.1016/j.phytol.2014.10.016
1
874-3900/ß 2014 Phytochemical Society of Europe. Published by Elsevier B.V. All rights reserved.

2
88
M. Ye et al. / Phytochemistry Letters 10 (2014) 287–290
Fig. 1. Chemical structures of steroid N-acetylglucosaminides 1–4.
3
Fig. 2. Observed key HMBC correlations of 1.
Moreover, a J HMBC correlation was observed from the
anomeric proton at 5.28 (1H, d, J = 7.3 Hz) to C-3 at 78.2,
demonstrating C-3 as the glycosidic linkage position (Fig. 2), while
the -glycosidic linkage could be easily determined based on the
d
d
Compound 2 was deduced to be a C-28 steroid N-acetylglu-
1
13
cosaminide by comparison of its H and C NMR data (Tables 1 and
) with those of compound 1. The obvious difference between 2
and 1 was that the terminal methyl at 0.89 (3H, t, J = 7.5 Hz, H
9) in 1 was replaced by an exomethylene group [ 4.87 and 4.85
each 1H, br s), 106.3 (CH ), 156.4 (qC)] in 2, which was supported
by the molecular formula (C36
exomethylene unit in the side chain of the aglycone in 2 was
confirmed by the HMBC correlations from H -26 ( 1.05)/H -27 (
.06) to C-24 ( 156.4). Thus, compound 2 was elucidated to be 3-
-O-(2-acetamido-2-deoxy- -glucopyranosyl)-ergosta-
,24(28)-diene.
Compounds 3 and 4 were obtained as a mixture, and both have
the same molecular formula (C36 ) based on their HRESIMS
C NMR spectroscopic data of 3 and 4
Tables 1 and 2) were quite similar to 1, indicating that 3 and 4
were also steroid N-acetylglucosaminide derivatives. 3 and 4
possessed the same aglycone of ergosta-5-en-3 -ol by comparison
b
2
observed coupling constant (7.3 Hz) of the anomeric proton. The
measured optical rotation value of the glucosamine hydrochlo-
d
3
-
2
0
2
(
d
ride, [
reported in the literature {[
Armarego and Chai, 2003). Finally, the structure of 1 was
elucidated to be 3- -O-(2-acetamido-2-deoxy- -glucopyrano-
syl)- -sitosterol. Actually, compound 1 has been previously semi-
synthesized by coupling of -sitosterol with glucosamine
a
]
D
2
+ 67.4 (c 0.05, H O), was in good accordance with that
2
5
d
2
a
]
D
2
+ 71.8 (after 20 h, c 4, H O)}
2
4(28)
6
H59NO .) of 2. The D
(
b
b-D
d
b
3
3
d
1
b
5
d
b
b-D
hydrochloride in 2009 (Grover et al., 2009). Herein, it was
reported as a new naturally occurring steroid N-acetylglucosa-
minide, and its proton and carbon NMR data were completely
assigned for the first time.
H61NO
6
1
13
data. The H and
(
Table 1
1
3
C NMR data of compounds 1–4 (125 MHz, C
5 5
D N).
b
13
No.
1
2
3
4
of its NMR spectroscopic data with those of 1. In the C NMR
spectrum of the mixture, two sets of carbon signals assignable to C-
17–C-24 (Table 1) were clearly observed, implying 3 and 4 were a
pair of C-24 epimers. As summarized by Wright et al. (Wright et al.,
978), for some diastereomeric C-24 alkyl sterols, slight differ-
ences in the chemical shifts of side-chain carbons permitted the
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
37.1
29.9
78.2
39.2
37.1
29.9
78.2
39.2
36.9
31.5
78.0
38.9
140.6
121.3
31.6
31.6
49.7
36.3
20.7
39.3
41.9
56.2
23.9
27.8
1
140.6
121.6
31.8
31.7
50.0
36.5
20.9
39.6
42.1
56.5
24.2
29.6
55.9
11.6
19.1
36.0
18.7
33.9
26.1
45.7
29.1
18.9
19.6
23.0
11.8
23.2
140.9
122.7
31.8
31.7
50.0
36.6
20.9
39.6
42.2
56.5
24.1
29.6
55.8
11.6
19.1
35.6
18.5
34.7
30.9
156.4
33.7
21.8
21.6
106.3
determination of the absolute configuration at C-24. As for 3 (24R),
C-26 (
only 1.8 ppm in contrast to a bigger difference of 3.0 ppm (C-26:
7.7; C-27: 20.7) in 4 (24S). Consequently, 3 and 4 were deduced as
(24R)-3- -O-(2-acetamido-2-deoxy- -glucopyranosyl)-
ergosta-5-ene and (24S)-3- -O-(2-acetamido-2-deoxy-
pyranosyl)-ergosta-5-ene, respectively.
To the best of our knowledge, none steroidal glycoside with a 2-
acetamido-2-deoxy- -glucose moiety has been so far isolated
d 20.1) and C-27 (d 18.3) have a chemical shift difference of
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
1
b
b-D
b
b-D-gluco-
b
-D
55.7
55.6
11.4
19.7
from vascular plants. Steroid N-acetylglucosaminides 1–4 were
tested for their cytotoxic effects against five human cancer cell
lines (BGC-823 and KE-97 gastric carcinoma, Huh-7 hepatocarci-
noma, Jurkat T cell lymphoblasts, and MCF-7 breast adenocarci-
35.5
18.8
33.4
29.6
38.5
32.0
20.1
18.3
14.9
35.8
18.6
TM
30.0
38.7
31.5
17.7
20.7
15.0
noma) using the CellTiter Glo luminescent cell viability assay (Ye
et al., 2013, 2014; Zang et al., 2013), but none of them was active
(
IC50s > 10
mM).
3. Experimental
OAc
23.3
170.3
100.5
57.8
23.0
170.0
100.2
57.7
75.8
71.9
78.2
62.2
3
.1. General procedures
1
70.3
0
1
100.5
57.8
76.1
72.2
78.2
62.5
0
Optical rotations were measured on a Perkin-Elmer 341 polar-
2
0
3
76.1
imeter (Perkin-Elmer, Waltham, MA, USA). IR spectra were
measured on an Avatar 360 FT-IR spectrophotometer (Thermo
Scientific, Waltham, MA, USA). NMR spectra were recorded on a
Bruker Avance DRX-500 spectrometer (Bruker Daltonics, Boston,
0
4
72.2
0
5
78.3
0
6
62.6

M. Ye et al. / Phytochemistry Letters 10 (2014) 287–290
289
Table 2
1
5 5
H NMR data of compounds 1–4 (500 MHz, J in Hz, C D N).
No.
1
1
2
3/4
1.68, 1H, m
1.64, 1H, m
1.69, 1H, m
0
.95, 1H, m
2.08, 1H, m
.73, 1H, m
0.91, 1H, m
0.83–0.80, 1H, m
2.08, 1H, m
2
2.06, 1H, m
1
1.73, 1H, m
1.74, 1H, m
3
4
3.87, 1H, dddd, 10.6, 9.4, 4.5, 3.9
2.67, 1H, m
3.88, 1H, dddd, 10.5, 10.2, 4.5, 3.9
2.67, 1H, m
3.87, 1H, dddd, 10.6, 9.4, 4.5, 3.9
2.66, 1H, m
2.50, 1H, m
2.42, 1H, m
2.50, 1H, m
6
7
5.33, 1H, br s
1.91, 1H, m
5.32, 1H, br s
1.91, 1H, m
5.32, 1H, br s
1.91, 1H, m
1.51, 1H, m
1.52, 1H, m
1.51, 1H, m
8
9
1.39, 1H, m
0.88, 1H, m
1.41, 2H, m
1.99, 1H, m
1.37, 1H, m
1.51–1.40, 1H, m
0.84, 1H, m
0.86, 1H, m
1
1
2
1.38, 2H, m
1.51, 2H, m
1
1.89, 1H, m
1.94–1.98, 1H, m
1.08, 1H, m
1.09, 1H, m
1.03, 1H, m
1
1
4
5
0.93, 1H, m
1.55, 1H, m
0.91, 1H, m
0.89, 1H, m
1.55, 1H, m
1.55, 1H, m
1.03, 1H, m
1.03, 1H, m
1.03, 1H, m
1
6
1.30, 2H, m
1.47, 1H, m
1.36–1.25, 2H, m
1.29, 1H, m
17
18
19
1.09, 1H, m
0.66, 3H, s
0.94, 3H, s
1.08, 1H, m
0.64, 3H, s
0.94, 3H, s
1.08, 1H, m
0.65, 3H, s
3: 0.85, 3H, s
4
: 0.87, 3H, s
2
0
1
1.39, 1H, m
1.41, 1H, m
1.51–1.40, 1H, m
2
0.99, 3H, d, 6.4
0.98, 3H, d, 5.5
3: 0.94, 3H, d, 6.8
4
: 0.96, 3H, d, 6.8
2
2
3
1.39, 1H, m
1.62, 1H, m
1.23, 1H, m
2.19, 1H, m
1.51–1.40, 1H, m
1.08, 1H, m
1.08, 1H, m
2
1.24, 2H, m
1.36–1.25, 2H, m
1.99, 1H, m
24
25
26
0.99, 1H, m
0.96, 1H, m
1.72, 1H, m
2.26, 1H, m
1.69, 1H, m
0.86, 3H, d, 6.9
1.05, 3H, d, 6.6
3: 0.83, 3H, overlapped
4
: 0.84, 3H, overlapped
3: 0.82, 3H, overlapped
: 0.81, 3H, overlapped
2
2
2
7
8
9
0.87, 3H, d, 5.4
1.26, 2H, m
1.06, 3H, d, 6.6
4
4.87, 1H, br s
3: 0.81, 3H, overlapped
4: 0.82, 3H, overlapped
4.85, 1H, br s
0.89, 3H, t, 7.5
COCH
NH
3
2.15, 3H, s
2.15, 3H, s
2.15, 3H, s
8.91, 1H, d, 5.8
5.28, 1H, d, 7.3
4.50, 1H, overlapped
4.50, 1H, overlapped
4.23, 1H, dd, 9.0, 7.1
3.98, 1H, m
8.91, 1H, d, 7.3
8.91, 1H, d, 6.7
0
1
5.28, 1H, d, 7.3
5.27, 1H, d, 7.5
0
2
4.50, 1H, overlapped
4.50, 1H, overlapped
4.23, 1H, dd, 9.4, 7.7
3.98, 1H, m
4.50, 1H, overlapped
4.50, 1H, overlapped
4.24, 1H, dd, 9.0, 7.4
3.98, 1H, m
0
3
0
4
0
5
0
6
4.56, 1H, br d, 11.6
4.56, 1H, br d, 11.4
4.38, 1H, dd, 11.4, 5.5
4.56, 1H, br d, 11.6
4.39, 1H, dd, 11.8, 5.3
4.38, 1H, dd, 11.7, 5.3
MA, USA). Chemical shifts are expressed in
d
(ppm) and are
August 2009, from Bozhou City, Anhui Province of China. The plant
was identified by Prof. Bao-Kang Huang (Department of Pharma-
cognosy, Second Military Medical University of China). A voucher
specimen (No. 100126) was deposited at the Herbarium of the
Department of Natural Products Chemistry, School of Pharmacy,
Fudan University.
referenced to the residual solvent signals. ESIMS were recorded on a
Bruker Daltonics micrOTOF-QII mass spectrometer (Bruker Dal-
tonics, Boston, MA, USA). EIMS were recorded on an Agilent 5975C
MSD mass spectrometer (Agilent Technologies, Santa Clara, CA,
USA). Semi-preparative HPLC was performed on a Beckman System
consisting of a Beckman Coulter System Gold 508 autosampler, Gold
1
26 gradient HPLC pumps with a Beckman System Gold 168 diode
3.3. Extraction and isolation
array detector (DAD) and a Sedex 80 (SEDERE, France) evaporative
light-scattering detector (ELSD) and a YMC-Pack ODS-A column
Dried and powdered fruits of L. japonicus (20.0 kg) were
extracted with MeOH (15 L ꢀ 4) under reflux for 4 h. The solvent
was removed at reduced pressure to give a dark brown residue
(
250 ꢀ 10 mm, dp 5
out on silica gel (200–300 mesh, Yantai Kang-Bi-Nuo Silysia
Chemical Ltd., Yantai, China), and TLC on silica gel (GF254
.25 mm, Yantai Kang-Bi-Nuo Silysia Chemical Ltd., Yantai, China).
SpotswerevisualizedusingUVlight (254 nm)and 15%H SO –EtOH.
mm). Column chromatography (CC) was carried
,
2
(ca. 832.6 g), which was then suspended in H O (2 L) and
0
extracted with EtOAc three times (3ꢀ 2 L). The entire EtOAc-
soluble extract (ca. 340.2 g) was then subjected to a flash CC over
silica gel eluting with gradients of petroleum ether (PE)–EtOAc–
MeOH (2:1:0 to 0:0:1, v/v/v) to yield ten fractions (Fr. 1–Fr. 10).
In an earlier work (Ye et al., 2014), Fr. 3 (PE-EtOAc, 1:1, v/v),
Fr. 4 (neat EtOAc), and Fr. 5 (EtOAc–MeOH, 20:1, v/v) were
found to yield a number of 28-nor-oleanane-type spirocyclic
2
4
3.2. Plant material
The dried fruits of L. japonicus were purchased from Shanghai
Jiu-Zhou-Tong Medicine Co. Ltd., and were originally collected in

2
90
M. Ye et al. / Phytochemistry Letters 10 (2014) 287–290
triterpenoids with interesting cytotoxic activity (Ye et al., 2014).
Only Fr. 8 (EtOAc–MeOH, 10:1, v/v, 28.2 g) was positive to
Liebermann–Burchard and Molish tests. The rest fractions were
not interesting and were discarded. Fr. 8 was later subjected to
Acknowledgements
This work was supported by NSFC grants (Nos. 81273401,
8
1202420, 21472021), grants from the Ph.D. Programs Foundation
of Ministry of Education (MOE) of China (Nos. 20120071110049,
0120071120049).
silica gel CC and eluted with EtOAc–EtOH–H
2
O (7:1:0.12, v/v/v)
and then separated by semi-preparative HPLC [96% MeOH in H
2
O
2
(
(
with 0.1% TFA), flow rate: 3.0 mL/min] to afford compounds 1
15.3 mg, t
R
= 37.1 min), 2 (2.2 mg, t
= 35.8 min).
R
= 27.5 min), and a mixture
of 3/4 (10.7 mg, t
R
Appendix A. Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.phytol.2014.10.016.
3
.3.1. Leonuroside A (1)
White amorphous powder; [
KBr) nmax: 3419, 3005, 2898, 2360, 1668, 1643, 1545, 1386, 1051,
22
a]
D
– 31.1 (c 0.22, MeOH); IR
(
6
ꢁ
1
1
13
73 cm ; H and C NMR data (in C
5
D
5
N) see Tables 1 and 2; (+)-
References
+
+
ESIMS: m/z 640 [M+Na] ; (+)-HRESIMS: m/z 640.4556 [M+Na]
calcd for C37 63NNaO , 640.4548).
Armarego, W.L.F., Chai, C.L.L., 2003. Purification of Laboratory Chemicals, fifth ed.
Butterworth-Heinemann, Oxford, pp. 251.
(
H
6
Ahmad, F., Ali, M., Alam, P., 2010. New phytoconstituents from the stem bark of
Tinospora cordifolia Miers. Nat. Prod. Res. 24, 926–934.
Gulavita, N.K., Wright, A.E., Kelly-Borges, M., Longley, R.E., Yarwood, D., Sills, M.A.,
3
.3.2. Leonuroside B (2)
White amorphous powder; [
KBr) max: 3420, 3012, 2975, 2360, 1680, 1669, 1351, 673 cm
22
a
]
D
– 68.2 (c 0.15, MeOH); IR
1994. Eryloside E from an Atlantic sponge Erylus goffrilleri. Tetrahedron Lett. 35,
ꢁ1
(
n
;
4299–4302.
1
13
Gutierrez-Lugo, M.T., Deschamps, J.D., Holman, T.R., Suarez, E., Timmermann, B.N.,
2004. Lipoxygenase inhibition by anadanthoflavone, a new flavonoid from the
aerial parts of Anadenanthera colubrina. Planta Med. 70, 263–265.
5 5
H and C NMR data (in C D N) see Tables 1 and 2; (+) ESIMS: m/z
+
+
6
C
24 [M+Na] ; (+)-HRESIMS: m/z 624.4236 [M+Na] (calcd for
59NNaO , 624.4235).
36
H
6
Grover, P., Singh, J., Dhar, K.L., Mehta, L., 2009. Synthetic modification of
b
-sitosterol
with 2-amino-1,3,4,6-tetra-O-hydroxy-2-deoxy-b-D-glucopyranose hydro-
chloride. J. Indian Chem. Soc. 86, 1112–1114.
3
.3.3. Leonurosides C (3) and D (4)
White amorphous powder; [
KBr) max: 3419, 3001, 2898, 2360, 1686, 1515, 1386, 1051,
Lin, A.S., Engel, S., Smith, B.A., Fairchild, C.R., Aalbersberg, W., Hay, M.E., Kubanek, J.,
2010. Structure and biological evaluation of novel cytotoxic sterol glycosides
from the marine red alga Peyssonnelia sp. Bioorg. Med. Chem. 18, 8264–8269.
Iida, T., Kasuga, Y., Arakawa, M., Mushiake, K., Ikegawa, S., Goto, J., Nambara, T.,
22
a
]
D
– 18.1 (c 0.08, MeOH); IR
(
6
n
ꢁ
1
1
13
12 cm ; H and C NMR data (in C
5
D
5
N) see Tables 1 and 2; (+)
2001. Complete
1
H and 13C resonance assignments of glycosidic conjugates of
+
+
ESIMS: m/z 626 [M+Na] ; (+)-HRESIMS: m/z 626.4436 [M+Na]
calcd for C36 61NNaO , 626.4391).
non-amidated and amidated bile acids. Magn. Reson. Chem. 39, 749–756.
Marschall, H.U., Egestad, B., Matern, H., Matern, S., Sj o¨ vall, J., 1989. N-acetylgluco-
saminides. A new type of bile acid conjugate in man. J. Biol. Chem. 264,
(
H
6
12989–12993.
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1.5 mL) was treated with 1.4 N HCl (5.0 mL), and refluxed at 85 8C
for 8 h. After neutralized with 0.7 M Na CO (5.0 mL), the solution
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phase was combined and then evaporated under vacuum to give
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(
2
3
Tachibana, K., Sakaitani, M., Nakanishi, K., 1985. Pavoninins, shark-repelling
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2
0:1) to furnish the aglycone (1a, 5.3 mg).
b
-Sitosterol (1a):
2
0
[
[
a
a
]
]
D
D
– 32.6 (c 0.2, CHCl
3
– 28.4 (c 0.1, CHCl )}; H and C NMR data (in CDCl ) were
3
) {lit. (Gutierrez-Lugo et al., 2004):
2
0
1
13
3
13
in full agreement with those reported in the literature (Ahmad
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+
3
96 (9), 381 (6), 273 (5), 232 (2), 231 (3), 213 (7), 43 (100).
Meanwhile, the aqueous layer was concentrated under reduced
pressure and then worked up as usual to give -glucosamine
O), lit. (Armarego and Chai,
D
2
0
hydrochloride {[
2
a
]
D
+ 67.4 (c 0.05, H
2
2
5
003): [
a
]
D
+ 71.8 (c 4, H O)}.
2
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Supporting information
Zang, Y., Xiong, J., Zhai, W.Z., Cao, L., Zhang, S.P., Tang, Y., Wang, J., Su, J.J., Yang, G.X.,
Zhao, Y., Fan, H., Xia, G., Wang, C.G., Hu, J.F., 2013. Fomentarols A–D, sterols from
the polypore macrofungus Fomes fomentarius. Phytochemistry 92, 137–145.
Supporting information (The 1D, 2D NMR and HR-ESIMS
spectra of compounds 1–4) of this article can be found online.