Total synthesis of α-galactosyl cerebroside

Article abstract of DOI:10.1016/S0008-6215(00)00092-6

A highly convergent synthetic approach was developed to obtain α- galactosyl cerebroside O-(α-D-galactopyranosyl)-2-hexacosylamino-D-ribo- 1,3,4-octadecantriol, which has previously been demonstrated to have immunostimulatory activity. Known 4,6-O-benzyli

Full text of DOI:10.1016/S0008-6215(00)00092-6

www.elsevier.nl/locate/carres
Carbohydrate Research 328 (2000) 95–102
Total synthesis of a-galactosyl cerebroside
Santiago Figueroa-Pe´rez, Richard R. Schmidt *
Fakulta¨t Chemie, Uni6ersita¨t Konstanz, Fach M 725, D-78457, Konstanz, Germany
Received 31 January 2000; accepted 13 March 2000
Abstract
A highly convergent synthetic approach was developed to obtain a-galactosyl cerebroside O-(a-
nosyl)-2-hexacosylamino- -ribo-1,3,4-octadecantriol, which has previously been demonstrated to have immunostimu-
latory activity. Known 4,6-O-benzylidene galactose was the starting material for both the required a-galactosyl and
the phytosphingosine building blocks O-(2,3-di-O-benzyl-4,6-O-benzylidene- -galactopyranosyl) trichloroacetimi-
date (4) and 2-O-methanesulfonyl- -arabino-1,2,3,4-octadecantetrol (5). The key step of the synthetic strategy is the
D-galactopyra-
D
D
D
highly regio- and stereoselective O-galactosylation of 1,3,4-O-unprotected phytosphingosine acceptor 5 using known
4 as donor. The total synthesis required only 11 synthetic steps starting from galactose. © 2000 Elsevier Science Ltd.
All rights reserved.
Keywords: Glycosphingolipids; Phytosphingosine; Synthesis; Glycosylation; Trichloroacetimidates
1. Introduction
cancer cells [6], foetus [7] and specific kidney
and intestine cell lines [8]. Owing to several
structural complexities of the natural product,
Koezuka and co-workers [9] have synthesized
several analogs for a structure–activity rela-
tionship study, taking the more prominent
compound agelasphin-9b {(2S,3S,4R)-1-O-
Monoglycosylated ceramides (cerebrosides)
are the simplest class of glycosphingolipids;
they are important surface molecules found in
virtually all cells. Galactosyl ceramides and
their metabolites have been shown to possess
important functions in promoting the regula-
tion of nerve cells [1], regulating protein ki-
nase C activities [2], and modulating the
function of hormone receptors [3].
Some a-galactosyl ceramides with phy-
tosphingosine structures, isolated from the
marine sponge Agelas mauritianus, were found
to have strong in vivo antitumor activities
against several murine tumor cells [4]. This
kind of structure has hardly been detected in
normal mammalian tissues [5], but mainly in
(a- -galactopyranosyl)-16-methyl-2[N-(R)-2-
D
hydroxytetracarbanoyl - amino] - 1,3,4 - hepta-
decanetriol} a lead structure, from which
compound 1 arose as a potent candidate for
clinical trial.
Compound 1 was also found to have im-
munostimulatory activity, since it is the ligand
of a T-cell antigen receptor expressed by Va 14
natural killer T-lymphocytes (NKT) [10].
NKT cells are known to play important roles
in the regulation of the progress of several
autoimmune diseases [11] and, when activated
with compound 1 and interleukin 12, they are
able to suppress tumor metastases [12].
These findings have promoted an increasing
demand for this compound for biological in-
* Corresponding author. Tel.: +49-7531-882538; fax: +
49-7531-883135.
E-mail address: rrs@dg3.chemie.uni-konstanz.de (R.R.
Schmidt).
0008-6215/00/$ - see front matter © 2000 Elsevier Science Ltd. All rights reserved.
PII: S0008-6215(00)00092-6

96
S. Figueroa-Pe´rez, R.R. Schmidt / Carbohydrate Research 328 (2000) 95–102
vestigations. Larger amounts of pure 1 are
therefore highly desirable. We report here on
a facile and convergent approach for the syn-
thesis of cerebroside 1, which can be easily
scaled up.
sequence is the regioselective a-galactosylation
of a 1,3,4-O-unprotected phytosphingosine
derivative (5 or 6), thus minimizing the protec-
tion–deprotection steps, which would make
the sequence shorter and therefore more effi-
cient. Trichloroacetimidate 4 can be readily
obtained from the known 4,6-O-benzylidene-
D
-galactose (7) [13], which via 8 should also
2. Results and discussion
provide 5 and 6.
The galactosyl donor should be designed in
order to obtain a high a:b ratio, thus easing
the purification of the resulting a-glycoside.
Generally, the 2,3,4,6-tetra-O-benzyl-galacto-
syl donor has been the choice for this case, but
it often led to anomeric mixtures that some-
times were difficult to purify. In our approach
we have placed a benzylidene group at C-4
The retrosynthetic analysis of compound 1
is depicted in Scheme 1. The first disconnec-
tion leads to fatty acid 3 and to the a-galacto-
sylphytosphingosine synthon 2, which in turn
has to be obtained starting from a suitable
protected galactosyl donor 4 and a phytosph-
ingosine acceptor bearing three chiral centers
in positions 2, 3 and 4. The key step of this
Scheme 1.

S. Figueroa-Pe´rez, R.R. Schmidt / Carbohydrate Research 328 (2000) 95–102
97
Scheme 2.
with those previously reported [15]. Com-
pound 9 was deallylated using PdCl2 without
affecting the benzylidene group in 74% yield
after purification by flash chromatography.
The analytical data of compound 9 were in
accordance with those already published
[15,16].
Reaction of the reducing sugar 10 with
trichloroacetonitrile and DBU in dry
dichloromethane afforded donor 4 in 77%
yield. The NMR data of this trichloroacetimi-
date were identical with those published by
Koeman et al. for the a anomer [17]. This
compound is relatively unstable although it
can be stored under argon for at least 3 weeks
at −18 °C.
and C-6, since there are examples in the litera-
ture that show that in the galactose series the
cis-decalin ring system with the equatorial
phenyl group hinders the attack from the b-
face [14]. Non-participating benzyl groups
were chosen to protect positions 2 and 3 of
the galactosyl trichloracetimidate 4. The phy-
tosphingosine triol acceptors (5 and 6) were
readily accessible from known 4,6-O-benzyli-
dene-
D
-galactose (7) [13].
-galactose (7) was 1-O-
4,6-O-Benzylidene-
D
allylated and 2,3-di-O-benzylated using a one-
pot procedure to afford an a/b mixture of
allyl 2,3-di-O-benzyl-4,6-O-benzylidene-galac-
topyranoside (9) in 69% yield (Scheme 2). The
NMR data of both compounds were identical

98
S. Figueroa-Pe´rez, R.R. Schmidt / Carbohydrate Research 328 (2000) 95–102
Acceptor 5 was obtained in 88% yield
catalysis to afford, after flash chromatogra-
phy, the a-glycoside 13 in 73% yield based on
trichloroacetimidate 4, and 83% based on the
consumed triol 5 (Scheme 3). Neither the b
anomer nor any other regioisomer was de-
through a controlled acid hydrolysis of acetal
12, which in turn was obtained from 4,6-O-
benzylidene- -galactose (7), via 8 and 11, as
D
previously described [18]. The multiplet at
1
1
4.72 ppm in the H NMR spectrum corre-
tected in the reaction. The H NMR spectrum
sponding to H-2 indicated the substitution of
this position by the methanesulfonyl (mesyl)
group, this was also confirmed by the presence
of a singlet at 3.14 ppm owing to the methyl
group of the mesylate. On the other hand,
acceptor 6 was prepared by azide substitution
of the mesylate group of 5, using tetra-
methylguanidinium azide as soluble azide ion
shows a doublet at 5.12 ppm with a coupling
constant of 3.54 Hz that corresponds to the
anomeric proton, it evidences a-stereochem-
13
istry of the glycosidic bond. In the C NMR
spectrum the signal of C-1 of the phytosphin-
gosine unit appears at 68.3 ppm (C-1% in 5:
59.9 ppm), which clearly indicated O-
alkylation.
When the same reaction was performed
with acceptor 6, the yield of pure compound 2
was only 45% based on 4 and 61% based on
reacted 6; obviously, the regioselectivity of the
galactosylation reaction is lower, thus result-
ing in a complex purification problem. The
presence of the more electron withdrawing
mesyl group seems to increase the reactivity
difference between primary and secondary hy-
droxyl groups.
1
donor. The multiplet of proton 2 in the H
NMR spectrum was strongly shifted upfield
(4.72“3.45 ppm) and the signal of carbon 2
13
in the C NMR was also shifted to 59.2 ppm,
evidencing the replacement of the mesylate
group by the azide group.
Regioselective galactosylation of compound
5 was performed with 0.75 equivalents of
trichloracetimidate 4 in THF at −10 °C, us-
ing trimethylsilyl trifluoromethanesulfonate
Scheme 3.

S. Figueroa-Pe´rez, R.R. Schmidt / Carbohydrate Research 328 (2000) 95–102
99
Table 1
Coupling constant values of H-2% in Hz
3. Experimental
General methods.—Solvents were purified in
the usual way. Thin-layer chromatography
(TLC) was performed on plastic plates Silica
J1%a-2%
J1%b-2%
J2%-3%
J3%-4%
13
2
5.6
2.8
4.4
3.4
B1
6.0
8.3
12.1
Gel 60 F254 and on HPTLC plates NH2 F254
S
(E. Merck, layer thickness 0.2 mm). The de-
tection was achieved by treatment with a solu-
tion of ammonium molybdate (20 g) and
cerium(IV) sulfate (0.4 g) in 400 mL of 10%
H2SO4 or with 15% H2SO4, and heating at
150 °C. Flash chromatography was carried
out on silica gel (Baker, 30–60 mm) and
Lichroprep NH2, particle size 40–63 mm (E.
Merck). Reversed-phase (RP) chromatogra-
phy: RP 18 (32–63 mm) ICN Biomedicals.
Medium-pressure liquid chromatography
(MPLC): LiChroprep Si 60 (E. Merck; size
15–25 mm), detection by differential refrac-
tometer. Optical rotations were determined at
25 °C with a Perkin–Elmer 241/MC polarime-
ter (1 dm cell). NMR spectra were recorded
with Bruker AC 250 and 600 DRX instru-
ments using tetramethylsilane as internal ref-
erence for CDCl3 and CD3OD, while for D2O
the water peak in 4.63 ppm was taken as the
The replacement of the mesylate group by
azide in compound 13 was performed by treat-
ment with five equivalents of tetra-
methylguanidinium azide in DMF at 40 °C for
2 days. Compound 2 was obtained in 89%
yield with complete inversion of configuration.
The 13C and 1H NMR signals for position 2 of
the phytosphingosine moiety were consider-
ably shifted (82.3“59.6 and 5.05“3.41
ppm), thus confirming the replacement of the
mesylate group by the azido group. All the
mesyl related signals disappeared. The dra-
matic change of the coupling constant pattern
of proton 2% (Table 1) confirms the inversion
of configuration at carbon 2%.
Compound 2 was submitted to hydrogenol-
ysis to remove the benzyl and benzylidene
groups and to reduce the azido group to the
amine group. The resulting compound was
isolated and without further purification
treated with hexacosanoic acid and the
reagent system N-hydroxybenzotriazole and
1 - [3 - (dimethylamino) - propyl] - 3 - ethylcarbo-
diimide hydrochloride in DMF at 40 °C. The
target compound 1 was obtained in pure form
after purification by RP column chromatogra-
phy and then by silica gel adsorption chro-
matography in 69% yield. All the analytical
data were in accord with those previously
published [9].
1
reference. The assignments of H NMR spec-
tra were based on chemical shift correlation
(DQF COSY) and rotating frame nuclear
Overhauser effect spectroscopy (ROESY). The
assignments of 13C NMR spectra were based
on carbon–proton shift-correlation heteronu-
clear multiple quantum coherence (HMQC).
MS spectra were recorded with MALDI-
Kompakt (Kratos), EI and FAB with Finni-
gan MAT 312/AMD equipment. Micro-
analyses were performed in the Microanalyses
Unit at the Fakulta¨t fu¨r Chemie, Universita¨t
Konstanz.
Allyl 2,3-di-O-benzyl-4,6-O-benzylidene- -
D
In conclusion, the total synthesis of com-
pound 1 was successfully achieved starting
galactopyranoside (9).—Sodium chloride (0.96
g, 22 mmol) was slowly added to a cooled
(ice-bath) solution of compound 7 [13] (5 g,
18.5 mmol) and allyl bromide (2.25 mL, 27.7
mmol) in dry DMF (70 mL). After the addi-
tion was completed, the mixture was stirred 30
min at room temperature (rt) and then diluted
with benzyl bromide (8.4 mL, 74 mmol). The
clear yellow solution was cooled again and
more NaH (1.92 g, 55.5 mmol) was supplied
in portions. The resulting suspension was fur-
from 4,6-O-benzylidene- -galactose (7), fol-
D
lowing a convergent approach, in 11 efficient
steps which involved a highly regio- and
stereoselective a-galactosylation of a 2-O-
methanesulfonyl -
D
- arabino - 1,2,3,4 - octade-
canetetrol acceptor. This facile sequence
permits the scaling up of this synthesis in
order to obtain bulk quantities of 1 for bio-
logical studies.

100
S. Figueroa-Pe´rez, R.R. Schmidt / Carbohydrate Research 328 (2000) 95–102
ther stirred at rt for 1 h, when a clear solution
was already formed. The reaction mixture was
then poured into ice water (1 L) and the
suspension was stirred until it reached rt. The
water was decanted and the residue was dis-
solved in CH2Cl2 (50 mL) and successively
washed with an aq 0.1 M HCl soln (20 mL),
aq satd NaHCO3 soln (15 mL), and water (15
mL), dried (MgSO4) and concentrated. Flash
chromatography of the residue (3:1 petroleum
ether–EtOAc) and recrystallisation from
EtOH provided compound 9 (a/b mixture) as
a white solid (6.28 g, 69%). TLC (3:1
petroleum ether–EtOAc): Rf 0.31, 0.38. The
NMR data of each anomer was identical, as
reported in the literature [15].
formed into a single slower moving spot (ꢀ3
h). The solution was then neutralized with
Et3N and concentrated to a thick syrup, which
was purified by flash chromatography (15:1
CH2Cl2–MeOH) to afford 5 (1.22 g, 88%) as
an amorphous solid. An analytical sample was
recrystallized from methanol: mp 129 °C; TLC
(3:1 petroleum ether–EtOAc): Rf 0.07 and
(10:1 CH2Cl2–MeOH): Rf 0.36; [h]D +18° (c
1
1, CHCl3); H NMR (600 MHz, Me2SO-d6): l
4.99 (s, 1 H, OH-1), 4.85 (d, 1 H, J3,OH 6.7 Hz,
OH-3), 4.72 (t, 1 H, J2,3=J1,2 5.7 Hz, H-2),
4.43 (d, 1 H, J4,OH 6 Hz, OH-4), 3.62 (m, 2 H,
H-1), 3.37 (m, 1 H, H-3), 3.33 (m, 1 H, H-4),
3.14 (s, 3 H, SCH3), 1.44 (m, 2 H, H-5), 1.25
(m, 24 H, CH2), 0.84 (t, 3 H, J 6.7 Hz, CH3).
13C NMR (150.9 MHz, Me2SO-d6): l 82.3
(C-2), 71.2 (C-3), 68.8 (C-4), 59.9 (C-1), 37.6
(SCH3), 33.1 (C-5), 31.2–22.0 (CH2), 13.8
(CH3). MALDIMS (positive mode, DHB/
THT-matrix): m/z 419.6 [MNa+], 435.6
[MK+]. Anal. Calcd for C19H40O6S (396.58):
C, 57.54; H, 10.16; S, 8.08. Found: C, 57.47;
H, 10.19.
2,3-Di-O-benzyl-4,6-O-benzylidene- -galac-
D
topyranose (10).—Palladium chloride (3.34 g,
21 mmol) was added to a solution of com-
pound 9 (1.5 g, 3.3 mmol) in 95% aq AcOH
(67 mL) containing AcONa (2.1 g). The dark
suspension was stirred until TLC evidenced
complete conversion of the reactand into a
slower moving spot. The reaction mixture was
filtered through a bed of Celite, diluted with
CH2Cl2 (100 mL), and washed with water (50
mL) satd NaHCO3 soln (2×30 mL) and wa-
ter (20 mL), dried (MgSO4) and concentrated.
The resulting dark yellow syrup was purified
by flash chromatography (2:1 petroleum
ether–EtOAc) to provide compound 10 (a/b
mixture) as a solid (1 g, 74%). The NMR data
of each anomer were identical with those re-
ported in the literature [15,16].
2-O-Azido- -ribo-1,2,3,4-octadecantetrol
D
(6).—Compound 5 (1.3 g, 3.27 mmol) was
stirred with tetramethylguanidinium azide (2.6
g, 16.39 mmol) in dry DMF (10 mL) at 50 °C
until TLC of the reaction mixture (10:1
CH2Cl2–MeOH) showed complete conversion
of the starting material into a faster moving
spot. The solution was then concentrated and
purified using silica gel flash column chro-
matography (30:1 CH2Cl2–MeOH) to afford
6 (1.22 g, 87%) as an amorphous solid. An
analytical sample was recrystallized from
MeOH: mp 90 °C; TLC (3:1 petroleum ether–
EtOAc): Rf 0.09 and (10:1 CH2Cl2–MeOH
O-(2,3-Di-O-benzyl-4,6-O-benzylidene- -
D
galactopyranosyl) trichloroacetimidate (4).—
To a solution of compound 10 (1 g, 2.4 mmol)
and CCl3CN (2.4 mL, 24 mmol) in dry
CH2Cl2 (20 mL) was added DBU (0.15 mL,
1.2 mmol) and the solution was stirred for 40
min at rt. The dark solution was then concen-
trated and purified by flash chromatography
(2:1 petroleum ether–EtOAc containing 0.1%
Et3N) to afford compound 4 (1.04 g, 77%) as
a white foam [17].
1
10:1): Rf 0.73; [h]D +10° (c 1, CHCl3); H
NMR (600 MHz, CDCl3): l 3.89 (m, 2 H,
H-1), 3.66 (m, 1 H, H-3%), 3.64 (m, 1 H, H-4%),
3.45 (m, 1 H, H-2%), 2.77 (d, 1 H, J3,OH 7.1 Hz,
OH-3%), 2.17 (d, 1 H, J4,OH 4.5 Hz, OH-4%),
1.50 (m, 2 H, H-5%), 1.26 (m, 24 H, CH2), 0.88
(t, 3 H, J 6.7 Hz, CH3). 13C NMR (150.9
MHz, CDCl3): l 74.3 (C-3%), 72.9 (C-4%), 61.6
(C-1%), 59.2 (C-2%), 33.4 (C-5%), 31.9–22.6
(CH2), 14.1 (CH3). MALDIMS (positive
mode, DHB/THT-matrix): m/z 366.5 [MNa+
], 382.5 [MK+]. Anal. Calcd for C18H37O3N3
(343.50): C, 62.93; H, 10.85; N, 12.23. Found:
C, 62.77; H, 10.99; N, 12.15.
2 - O - Methanesulfonyl -
D
- arabino - 1,2,3,4-
octadecantetrol (5).—To a solution of com-
pound 12 [18] (1.7 g, 3.49 mmol) in methanol
(50 mL) was added p-toluenesulfonic acid (40
mg), and the solution was stirred at rt until
the starting material was completely trans-

S. Figueroa-Pe´rez, R.R. Schmidt / Carbohydrate Research 328 (2000) 95–102
101
1-O-(2,3-Di-O-benzyl-4,6-O-benzylidene-h-
-galactopyranosyl)-2-O-methanesulfonyl-
mL) soln, water (7 mL), then dried (MgSO4),
filtered and concentrated. Flash chromatogra-
phy of the residue (3:1 petroleum ether–
EtOAc) and recrystallization (EtOAc–
pentane) provided 2 (0.64 g, 89%) as colorless
crystals. Method (b): to a solution of com-
pound 6 (0.51 g, 1.5 mmol) and compound 4
(0.42 g, 0.75 mmol) in dry THF (10 mL) was
added trimethylsilyl trifluoromethanosul-
fonate (13 mL, 0.075 mmol) at −20 °C, and
the solution was stirred for 30 min. The solu-
tion was then neutralized with Et3N and con-
centrated to dryness. Flash chromatography
of the residue (3:1–1:1 petroleum ether–
EtOAc) provided recovered 6 (0.32 g, 62% of
the starting material) and 2 (0.26 g, 49% based
on trichloroacetimidate 4, and 61% based on
consumed acceptor 6): mp 70.8 °C; TLC (1:1
petroleum ether–EtOAc): Rf 0.39. [h]D +65°
D
D-
arabino-1,2,3,4-octadecantetrol (13).—To a
solution of 5 (0.95 g, 2.4 mmol) and 4 (0.9 g,
1.6 mmol) in dry THF (20 mL) was added
trimethylsilyl trifluoromethanosulfonate (30
mL, 0.16 mmol) at −10 °C, and the solution
was stirred for 30 min. The solution was then
neutralized with Et3N and concentrated to
dryness. Flash chromatography of the residue
(3:1–1:1 petroleum ether–EtOAc) provided
recovered 5 (0.4 g, 41% of the starting
amount) and a-glycoside 13, which was recrys-
tallized from EtOAc–hexane (0.96 g, 73%
based on 4 and 83% based on reacted 5) as
colorless needles: mp 137.5 °C; TLC (1:1
petroleum ether–EtOAc): Rf 0.28; [h]D +62°
1
(c 1, CHCl3); H NMR (600 MHz, CDCl3): l
7.51–7.25 (m, 15 H, PhꢀH), 5.49 (s, 1 H, CH),
5.05 (m, 1 H, H-2%), 5.00 (d, 1 H, J1,2 3.8 Hz,
H-1), 4.77 (AB, J 11.4, CH2Ph-2), 4.74 (AB, J
12.8, CH2Ph-3), 4.23 (d, 1 H, J3,4 3.4 Hz,
J4,5B1 Hz, H-4), 4.22 (m, 1 H, H-6a), 4.09
(dd, 1 H, J2,3 10 Hz, H-2), 4.03 (dd, 1 H, J1%a,1%b
11.7 Hz, H-1%a), 4.01 (m, 1 H, H-6b), 3.86 (m,
1 H, H-3), 3.77 (dd, 1 H, H-1%b), 3.54 (m, 1 H,
H-4%), 3.49 (m, 1 H, H-3%), 2.96 (s, 3 H, SCH3),
2.66 (d, 1 H, J4,OH 5.6 Hz, OH-4%), 2.62 (d, 1
H, J3,OH 7.6 Hz, OH-3%), 1.52 (m, 2 H, H-5%),
1.27 (m, 24 H, CH2), 0.88 (t, 3 H, J 6.7 Hz,
CH3). 13C NMR (150.9 MHz, CDCl3): l
138.3, 138.0, 137.6 (ipso), 128.9–126.2 (Ph),
101.1 (CH), 99.3 (C-1), 79.1 (C-2%), 75.5 (C-3),
75.2 (C-2), 74.0 (C-4, C-3%), 73.8 (CH2Ph-2),
71.4 (CH2Ph-3), 70.5 (C-4%), 69.2 (C-6), 68.2
(C-1%), 63.1 (C-5), 38.4 (SCH3), 33.0 (C-5%),
31.9–22.6 (CH2), 14.1 (CH3). MALDIMS
(positive mode, DHB/THT-matrix): m/z 849.7
[MNa+]. Anal. Calcd for C46H66O11S (827.02):
C, 66.80; H, 8.04; S, 3.87. Found: C, 66.69; H,
8.12.
1
(c 1, CHCl3); H NMR (600 MHz, CDCl3): l
7.51–7.25 (m, 15 H, PhꢀH), 5.46 (s, 1 H, CH),
4.86 (d, 1 H, J1,2 3.6 Hz, H-1), 4.79 (AB, J
11.6, CH2Ph-2), 4.74 (d, J 12.0, CH2Ph-3),
4.23 (m, 1 H, J6a,6b 12.4, H-6a), 4.19 (d, 1 H,
J3,4 3.4 Hz, J4,5B1 Hz, H-4), 4.17 (dd, 1 H,
J1%a,1%b 10.6 Hz, H-1%a), 4.08 (dd, 1 H, J2,3 10
Hz, H-2), 4.02 (m, 1 H, J6b,5 1.2, H-6b), 3.97
(m, 1 H, H-3), 3.86 (dd, 1 H, H-1%b), 3.75 (m,
1 H, H-3%), 3.60 (m, 1 H, H-4%), 3.47 (d, 1 H,
J3,OH 7.1 Hz, OH-3%), 3.41 (m, 1 H, H-2%), 2.17
(d, 1 H, J4,OH 4.5 Hz, OH-4%), 1.50 (m, 2 H,
H-5%), 1.26 (m, 24 H, CH2), 0.88 (t, 3 H, J 6.7
Hz, CH3). 13C NMR (150.9 MHz, CDCl3): l
138.3, 137.8, 137.7 (ipso), 128.9–126.2 (Ph),
101.0 (CH), 99.9 (C-1), 76.3 (C-3), 74.8 (C-2),
74.7 (C-3%), 74.1 (C-4), 73.8 (CH2Ph-2), 71.4
(CH2Ph-3), 72.8 (C-4%), 69.6 (C-1%), 69.4 (C-6),
63.3 (C-5), 59.6 (C-2%), 32.4 (C-5%), 31.9–22.6
(CH2), 14.1 (CH3). MALDIMS (positive
mode, DHB/THT-matrix): m/z 796.9 [MNa+
]. Anal. Calcd for C45H63N3O8 (774.00): C,
69.83; H, 8.20; N, 5.42. Found: C, 69.67; H,
8.35; N, 5.38.
2-Azido-1-O-(2,3-di-O-benzyl-4,6-O-benzy-
lidene - h -
D
- galactopyranosyl) -
D
- ribo - 1,3,4-
octadecantriol (2).—Method (a): to a solution
of compound 13 (0.9 g, 1.08 mmol) in dry
DMF (5 mL) was added tetramethyl-
guanidinium azide (0.86 g, 5.44 mmol), and
the solution was stirred for 48 h at rt. The
mixture was then concentrated to dryness and
the resulting syrup dissolved in CH2Cl2 (20
mL). The solution was successively washed
with 0.03 M HCl (3×5 mL), satd NaHCO3 (7
1-O-(h-
D
-Galactopyranosyl)-2-hexacosyla-
mino- -ribo-1,3,4-octadecantriol (1).—A mix-
D
ture of compound 2 (123 mg, 0.15 mmol) and
Pd–C 10% (0.1 g) in MeOH (10 mL) contain-
ing AcOH (0.1 mL) was stirred under a H2
atmosphere at rt for 12 h. The mixture was
filtered, concentrated and coevaporated with
toluene. The resulting syrup was then

102
S. Figueroa-Pe´rez, R.R. Schmidt / Carbohydrate Research 328 (2000) 95–102
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dissolved in dry DMF (3 mL), hexacosanoic
acid (58 mg, 0.15 mmol), N-hydroxybenzotri-
azol (23 mg, 0.15 mmol) and 1-[3-(dimethy-
lamino) - propyl] - 3 - ethylcarbodiimide hydro-
chloride (32 mg, 0.15 mmol) were successively
added and the solution was stirred at 40 °C
for 12 h. The mixture was injected onto a RP
silica gel C-18 column and eluted with MeOH
containing increasing amounts of CH2Cl2, the
nearly pure white solid so obtained was
rechromatographed on a short silica gel
column (25:1 CH2Cl2 –MeOH) to provide 1
(94 mg, 69%) as an amorphous solid. The
NMR data were identical with those reported
in the literature [9]. TLC (15:1 CH2Cl2–
MeOH): Rf 0.49. Anal. Calcd for C50H99NO9
(858.33): C, 69.96; H, 11.62; N, 1.63. Found:
C, 69.88; H, 11.81; N, 1.54.
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(1995) 6839–6842.
Acknowledgements
This work was supported by the Deutsche
Forschungsgemeinschaft and the Fonds der
Chemischen Industrie. We are grateful to Dr
Armin Geyer for his help in the structural
assignments.
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