Structure determination and synthesis of a new cerebroside isolated from the traditional Chinese medicine Typhonium giganteum Engl.

Article abstract of DOI:10.1016/S0040-4039(02)00583-X

A new cerebroside with C18-4,8-sphingadienine as the long chain base has been isolated from the traditional Chinese medicine Typhonium giganteum Engl., and its structure was determined by 2D NMR and MS methods. It was then synthesized using D-xylose and a

Full text of DOI:10.1016/S0040-4039(02)00583-X

TETRAHEDRON
LETTERS
Pergamon
Tetrahedron Letters 43 (2002) 3529–3532
Structure determination and synthesis of a new cerebroside
isolated from the traditional Chinese medicine
Typhonium giganteum Engl.
Xuesong Chen,a,b Yu-Lin Wua,* and Dihua Chenb,*
aState Key Laboratory of Bio-organic and Natural Products Chemistry, Shanghai Institute of Organic Chemistry,
Chinese Academy of Sciences, 354 Fenglin Road, Shanghai 200032, China
bInstitute of Medicinal Plant Development, Chinese Academy of Medical Sciences & Peking Union Medical College,
100 Dongbeiwang Road, Beijing 100094, China
Received 14 December 2001; revised 12 March 2002; accepted 21 March 2002
Abstract—A new cerebroside with C18-4,8-sphingadienine as the long chain base has been isolated from the traditional Chinese
medicine Typhonium giganteum Engl., and its structure was determined by 2D NMR and MS methods. It was then synthesized
using
D-xylose and ascorbic acid as the chiral starting materials. © 2002 Elsevier Science Ltd. All rights reserved.
The dried root tuber of Typhonium giganteum Engl. is
recorded in Chinese pharmacopoeia as a traditional
Chinese medicine and named Baifuzi in Chinese.1 It has
the effect of ‘dispelling wind-phelgm’ and has been used
for the treatment of cerebral apoplexy for a long time
in China. However, the active components of the
medicine were not clear until now.
reviews on the biological activities of cerebrosides, sph-
ingosine and sphingolipids.3 A significant amount of
synthetic work4 on cerebrosides has been reported. Due
to their scarcity in natural sources and their interesting
biological activities, in addition to isolation and struc-
ture determination we also began total syntheses of
these new cerebrosides. Herein, we report the structure
determination and the synthesis of typhoniside A (Fig.
1), a representative of these cerebrosides.
In our research searching for the biologically active
constituents from T. giganteum Engl., we have isolated
eight cerebrosides for the first time from this plant. It
has been reported that cerebrosides act by signal trans-
duction through cell membranes and exhibit significant
activities. In 1991, Harouse2a reported that galactosyl
ceramide inhibited the entry of HIV-1 in neural cell
lines. In 1990, Kitagawa reported2b that cerebrosides
from soybean, the seeds of Glycine max Merrill, showed
ionophoric activity for Ca2+ ions. There are many
The EtOH extract (310 g) of T. giganteum Engl. (20 kg)
collected from Yu county Henan province, China, was
divided into three fractions using hexane, EtOAc and
95% EtOH. The hexane fraction (75 g) was subjected to
column chromatography over silica gel (eluent: P.E./
Me2CO/MeOH) to give eight crude fractions. Fraction
6 (2.8 g) was chromatographed using a silica gel column
(eluent: CHCl3/MeOH, 95:5) to afford 200 mg of a
Figure 1. The structure of typhoniside A (1).
Keywords: Typhonium giganteum Engl.; cerebroside; typhoniside.
* Corresponding authors. Fax: 86 10 67635913; e-mail: ylwu@pub.sioc.ac.cn
0040-4039/02/$ - see front matter © 2002 Elsevier Science Ltd. All rights reserved.
PII: S0040-4039(02)00583-X

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X. Chen et al. / Tetrahedron Letters 43 (2002) 3529–3532
mixture of cerebrosides, which was purified further to
give eight cerebrosides by HPLC (eluent: MeOH/H2O,
96:4) including typhoniside A (6.0 mg).
311.3314) and at m/z 306.2489 (calcd for C19H32O2N:
306.2433) were observed for part of the a-hydroxy-
docosanoyl acid and the fragment of the a-hydroxy-
docosanoyl acid due to a Mclafferty rearrangement.
The number of carbons in the LCB and FA were
determined to be 18 and 22, respectively.
Typhoniside A was a white amorphous powder with
mp 172–174°C. Its molecular formula was determined
as C46H87O9N by HRMS m/z 798.6451 [M+H]+ (calcd
for C46H88O9N m/z 798.6453). Its IR spectrum (3400,
The carbon chemical shifts at l 70.1 (C-1), 54.7 (C-2),
72.6 (C-3), 175.6 (C-1%) and 72.4 (C-2%) were in agree-
ment with those reported for other (2S,3R,2%R) sphin-
gosine moieties.6 Thus, the structure of 1 was assigned
1
1630, 1080, 720 cm−1) and H and 13C NMR spectra
(Table 1) suggested the glycosphingolipid nature.
as
1-O-b-D-glucopyranosyl-(2S,3R,4E,8E)-2-[2%(R)-
The signals at l 105.6, 78.5, 71.7, 78.5 and 62.8 in the
13C NMR spectrum suggested that the sugar moiety in
1 was a b-glucopyranoside. The coupling constant
between H-1%% [l 4.91 (1H, d, 7.7 Hz)] and H-2%% [l 4.03
hydroxydocosanoylamino]-4,8-octadecadiene-1,3-diol.
After the structure had been determined we designed a
facile and convergent approach for the synthesis of
typhoniside A to confirm the structure and try to find
another way to provide this cerebroside for biological
investigation.
(1H, t, 7.7 Hz)] supported the b-D-configuration of the
sugar. The 4,5 alkenyl bond was found to be trans, as
evidenced by the vicinal coupling constant (J4,5=15.3
Hz). The 8,9 alkenyl bond was also found to be trans,
as evidenced by the chemical shifts of C-7 and C-10
(32.88, 32.75). Usually the signals of the carbons next
to a trans double bond appear at l 32–33.5 With this
information we determined that 1 is a glucosylceramide.
Based on the retrosynthetic analysis depicted in Scheme
1, the molecule 1 was divided into three fragments. The
sphingadiene fragment 2 was synthesized from D-xylose
via a SN2% type of reaction mediated by a thioether
carbanion as in our previous report.7 The chirally pure
FA part 3, (R)-a-hydroxydocosanoyl acid, was pre-
pared from (R)-4-hydroxytetrahydrofuran-2-one 4,
The length of the long chain base (LCB) and the fatty
acid (FA) were determined by HREIMS. Two diagnos-
tic fragment ions at m/z 311.3259 (calcd for C21H43O:
which in turn could be obtained from L
-ascorbic acid.8
The synthesis from 4 to 3 is shown in Scheme 2.
1
Table 1. NMR data of compound 1 (500 MHz for H
and 125 MHz for 13C NMR in C5D5N)
Treatment of (R)-4-hydroxytetrahydrofuran-2-one 48
with a catalytic amount (20 mol%) of trimethylsilyl
trifluoromethanesulfonate (TMS-OTf) and 1.5 equiva-
lents of benzyl trichloroacetimidate prepared by sodium
hydride catalyzed addition of benzyl alcohol to
trichloroacetonitrile9 yielded 5, [h]2D0 +35.9 (c 0.6,
CH2Cl2). Lactone 5 was converted to hemiacetal 6 by
DIBAL-H reduction. The Wittig reagent generated
from the octadecanyl bromide phosphonium salt
reacted with hemiacetal 6 to give alcohol 7, which was
oxidized to afford acid 8. Removal of the benzyl pro-
tecting group and hydrogenation of the double bond
gave 3, [h]2D0 +12 (c 0.93, pyridine).
Position
1
1H NMR (J in Hz)
13C NMR
70.1
HMBC
4.70 (1H, dd, 10.3, 5.8)
4.25 (1H, dd, 10.3, 4.0)
4.81 (1H, m)
1%%
2
3
4
5
6
7
8
9
54.7
72.594a
132.112b
131.988b
32.93c
32.88c
129.96
131.14
32.75c
29.51–30.18
32.1
22.9
14.2
175.6
72.396a
35.7
25.9
29.5–30.2
32.1
22.9
14.2
105.6
75.1
78.5
71.7
78.5
1
4.76 (1H, t, 6.1)
5.93 (1H, dd, 15.3, 6.1)
6.00 (1H, dt, 15.3, 5.3)
2.16 (2H, s-like)
2.01 (2H, s-like)
5.49 (1H, s-like)
5.49 (1H, s-like)
2.16 (2H)
1, 2
3, 4
4, 6
5, 7
6, 8
7, 9
8, 10
9
10
11–15
16
17
18
1%
2%
3%
4%
5%–19%
20%
21%
22%
1%%
2%%
3%%
4%%
5%%
6%%
1.25–1.30 (10H)
1.25–1.30 (2H)
1.25–1.30 (2H)
Thereafter ceramide 12 was synthesized as shown in
Scheme 3. Acylation of 3 with Ac2O yielded 9, [h]D20
+6.6 (c 0.5, CHCl3).10 Acid 9 was treated N-hydroxy-
succinimide in the presence of DCC to give amide 10,
which was reacted with sphingadiene 2 in the presence
of DMAP and Et3N to yield 11, [h]2D0 +10.7 (c 0.5,
CHCl3). Removal of the protecting group with K2CO3
gave ceramide 12. The ceramide 12 was converted to
the glycosyl acceptor 13, [h]2D0 +6.6 (c 0.5, CHCl3), using
a conventional method10 in 77% overall yield.
0.86 (3H, t, 6.0)
4.57 (1H, dd, 7.7, 3.6)
2.04 (2H)
1.78 (2H)
1.25–1.30 (30H)
1.25–1.30 (2H)
1.25–1.30 (2H)
0.86 (3H, t, 6.0)
4.91 (1H, d, 7.7)
4.03 (1H, t, 7.7)
4.24 (1H, m)
4.24 (1H, m)
3.90 (1H, m)
4.52 (1H, dd, 12.0, 1.9)
4.34 (1H, dd, 12.0, 5.4)
8.35 (1H, d, 9)
1
1
3%%
4%%
5%%
6%%
5%%
The final glycosylation was performed in the usual way.
Thus ceramide 13 was treated with 2,3,4,6-tetra-O-ben-
zoyl-a-
-glucopyranosyl trichloroacetimedate 1411 in
D
the presence of a catalytic amount of TMSOTf (0.05
equiv.) in CH2Cl2 to provide the 1-O-glucosylated
product 15, [h]2D0 +20.8 (c 1.3, CHCl3). Deprotection of
the alcohol groups in 15 yielded 1, [h]2D0 +4.0 (c 0.28,
CHCl3/MeOH 1:1). All the spectroscopic properties of
62.8
N–H
1%
Note: a, b, c indicates that the chemical shifts are interchangeable.

X. Chen et al. / Tetrahedron Letters 43 (2002) 3529–3532
3531
Scheme 1. Retrosynthetic analysis of typhoniside A.
Scheme 2. Reagents and conditions: (a) TMSOTf, CCl3C(NH)OBn, cyclohexane: CH2Cl2 (2:1), 71%; (b) DIBAL-H, CH2Cl2, 74%;
(c) BuLi, C18H37PPh3Br, THF, 0°C, 95%; (d) PDC, DMF, rt 32 h, 68%; (e) 10% Pd/C, cyclohexane:EtOH (1:2), 40°C, 20 h, 88%.
Scheme 3. Reagents and conditions: (a) Ac2O, DMAP, CH2Cl2, overnight, pyridine, rt, 100%; (b) N-hydroxysuccinimide, DCC,
CH2Cl2, 5 h, 80%; (c) 2, DMAP, Et3N, THF, 8 h, 95%; (d) K2CO3, MeOH, 90%; (e) (i) TrCl, pyridine, DMAP, 80°C; (ii) BzCl,
pyridine, 14 h; (iii) p-TsOH, CH2Cl2:MeOH (2:1) 8 h, 77% over the three steps.
the product were in agreement with those of the iso-
lated natural material (Scheme 4).12
Acknowledgements
In conclusion, a new cerebroside, typhoniside A (1) has
been isolated from the traditional Chinese medicine
Typhonium giganteum Engl., and was synthesized via
the chiron approach.
We are grateful to the National Natural Science Foun-
dation of China (29790126), the Chinese Academy of
Sciences (95-A1-504-03) and the Ministry of Science
and Technology of China (970211006-6, G2000077502)

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X. Chen et al. / Tetrahedron Letters 43 (2002) 3529–3532
Scheme 4. Reagents and conditions: (a) TMSOTf, CH2Cl2, 65%; (b) NaOMe, MeOH, 80%.
for financial support. We also thank Professor Zhu-Jun
Yao, Professor Yikang Wu and Assoc. Professor Jian-
yong Si for helpful discussions.
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5. Stotuers, J. B. Carbon-13 NMR Spectroscopy; Academic
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0.15, CHCl3/MeOH, 1:1) H and 13C NMR, see Table 1,
HRMS (FAB, positive) calcd for C46H88O9N (M+H)
798.6453. Found: 798.6453.