De novo sythesis of a D-galacturonic acid thioglycoside as key to the total synthesis of a glycosphingolipid from

Article abstract of DOI:10.1021/ol800227b

A concise synthesis of a differentially protected D-galacturonic acid (D-GalA) thioglycoside and the construction of a potent immunomodulating glycosphingolipid are described. The key steps of the synthesis are an Evans aldol reaction between a C4 aldehyde and a PMB-protected glycolyloxazolidinone as well as a tandem-PMB-deprotection/cyclization to thioglycosides. The key glycosylation step is optimized by varying the anomeric leaving group, the activating agent, and the solvent system.

Full text of DOI:10.1021/ol800227b

ORGANIC
LETTERS
2008
Vol. 10, No. 8
1573-1576
De Novo Synthesis of a -Galacturonic
D
Acid Thioglycoside as Key to the Total
Synthesis of a Glycosphingolipid from
Sphingomonas yanoikuyae
Pierre Stallforth, Alexander Adibekian, and Peter H. Seeberger*
Laboratory for Organic Chemistry, Swiss Federal Institute of Technology (ETH)
Zurich, Wolfgang-Pauli-Strasse 10, HCI F312 CH-8093 Zurich, Switzerland
seeberger@org.chem.ethz.ch
Received February 4, 2008
ABSTRACT
A concise synthesis of a differentially protected D-galacturonic acid (D-GalA) thioglycoside and the construction of a potent immunomodulating
glycosphingolipid are described. The key steps of the synthesis are an Evans aldol reaction between a C4 aldehyde and a PMB-protected
glycolyloxazolidinone as well as a tandem-PMB-deprotection/cyclization to thioglycosides. The key glycosylation step is optimized by varying
the anomeric leaving group, the activating agent, and the solvent system.
D-Galacturonic acid (D-GalA) constitutes the most abundant,
naturally occurring uronic acid. As a R-(1-4)-linked ho-
mopolymer, homogalacturonane, it forms the major constitu-
ent of pectin,1 and as a R-(1-4)-linked species it is also
present in the immunogenic lipopolysaccharides of various
pathogenic bacteria.2 Furthermore, several bacterial gly-
cosphingolipids contain D-GalA that forms an R-glycosidic
linkage to a ceramide (e.g., A, Scheme 1).3 These glycolipids
exhibit an intriguing biological activity, since they are
recognized by human natural killer T (NKT) cells, after
binding to CD1d, and induce immune response.4 NKT cell
activation has been studied in the past decade in the context
of autoimmune diseases and cancer research.5 Efficient
syntheses of R-(1-4)-linked D-GalA containing biologically
active molecules as molecular probes for biomedical inves-
tigations are thus needed. Although oligosaccharide synthesis
has been subjected to a large number of improvements, a
typical monosaccharide building block synthesis still mainly
relies on tedious protection and deprotection steps in order
to access the required protecting group pattern.6 De novo
synthesis7 circumvents these disadvantages by the use of
preprotected linear fragments and not starting from the
corresponding unprotected sugar.
Here, we report a short and high yielding synthesis of
differentially protected D-galacturonic acid thioglycosides as
part of our ongoing program directed at the de novo synthesis
of selectively protected carbohydrate building blocks.8 Our
building block was then used to synthesize a highly im-
(1) Willats, W. G. T.; McCartney, L.; Mackie, W.; Knox, J. P. Plant
Mol. Biol. 2001, 47, 9.
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D.; Tsuji, M.; Kawahara, K.; Wong, C.-H.; Kronenberg, M. Nature 2005,
434, 520. (b) Wu, D.; Xing, G. W.; Poles, M. A.; Horowitz, A.; Kinjo, Y.;
Sullivan, B.; Bodmer-Narkevitch, V.; Plettenburg, O.; Kronenberg, M.;
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10.1021/ol800227b CCC: $40.75
© 2008 American Chemical Society
Published on Web 03/26/2008

following two protection-deprotection steps. The linear C6-
acetal C can be dissected into monoprotected C4-aldehyde
E and known glycolate oxazolidinone D12 by a retroaldol
reaction. In order to establish the desired configuration on
C-4 and C-5, a 2,3-syn-3,4-anti-selective Evans aldol
reaction13-15 is planned.
Scheme 1. Retrosynthesis of Glycosphingolipid A
The doubly benzylated aldehyde E can be readily obtained
from commercially available L-arabinose ethyl dithioacetal
in four steps.8a
Our synthetic efforts started with trials focusing on the
2,3-syn-3,4-anti-selective Evans aldol reaction as the key step
in the building block synthesis (Table 1). In principle, both
Table 1. Evans Aldol Reaction
munogenic galacturonosylceramide (A) from Sphingomonas
yanoikuyae.9
entry donor
conditions
dr
yield (%)
The retrosynthetic plan is presented in Scheme 1. Glyco-
conjugate A can be dissected into benzyl-protected ceramide
F and D-GalA building block B. For the synthesis of the
ceramide part, sphingosine G and fatty acid H are connected
by amide coupling. Acid H is derived from 1-tetradecene
via a dihydroxylation/oxidation sequence, whereas amino
alcohol G is prepared according to Howell et al.10 starting
from the Weinreb amide of N-Boc-L-serine.11 Key thiogly-
coside B will be formed by cyclization of thioacetal C,
1
2
3
4
5
6
7
D1
D1
D1
D2
D2
D2
D2
Bu2BOTf, NEt3, toluenea,14
TiCl4, DIPEA, CH2Cl2, 0 °C15
np
np
np
np
49
50
90
b,14
TiCl4, DIPEA, CH2Cl2
b
LDA, CH2Cl2
LDA, tolueneb
LDA, Et2Ob
LDA, THFb
2.3:1
4.6:1
4.9:1
a Temperature: -50 to -30 °C. b -78 °C.
(6) (a) Paulsen, H. Angew. Chem., Int. Ed. Engl. 1982, 21, 155. (b)
Danishefsky, S. J.; McClure, K. F.; Randolph, J. T.; Ruggeri, R. B. Science
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B.; Ranzinger, R.; Herget, S.; Adibekian, A.; von der Lieth, C.-W.;
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1993, 3, 97. (h) Roy, R. Drug DiscoVery Today, 2004, 1, 327. (i) Dube, D.
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J. AdV. Carbohydr. Chem. Biochem. 2004, 59, 69. (k) Dwek, R. A. Chem.
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Dinkelaar, J.; Overkleeft, H. S.; van der Marel, G. A. Eur. J. Biochem.
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(7) For recent organocatalytic de novo syntheses of partially protected
monosaccharides, see: (a) Northrup, A. B.; MacMillan, D. W. C. Science
2004, 305, 1752. (b) Enders, D.; Grondal, C. Angew. Chem. 2005, 117,
1235; Angew. Chem., Int. Ed. 2005, 44, 1210. (c) Utsumi, N.; Imai, M.;
Tanaka, F.; Ramasastry, S. S. V.; Barbas, C. F., III. Org. Lett. 2007, 9,
3445. For reviews on the de novo synthesis of carbohydrates, see: (d)
Schmidt, R. R. Pure Appl. Chem. 1987, 59, 415. (e) Kirschning, A.;
Jesberger, M.; Schoning, K.-U. Synthesis 2001, 507. (f) Vogel, P. In The
Organic Chemistry of Sugars; Levy, D. E., Fugedi, P., Eds.; Taylor and
Francis Group/CRC Press: Boca Raton, 2006; Chapter 13, p629.
(8) (a) Timmer, M. S. M.; Adibekian, A.; Seeberger, P. H. Angew. Chem.,
Int. Ed. 2005, 44, 7605. (b) Adibekian, A.; Bindschadler, P.; Timmer, M.
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13, 4510.
enantiomeric forms of the oxazolidinone auxiliary can be
employed to construct the desired diastereomer.13 The desired
aldol product was obtained after considerable trials by an
LDA-promoted Evans aldol reaction at -78 °C with auxiliary
D211 in toluene (Table 1, entry 7). Aldol 2 was obtained in
a 90% yield and 4.9:1 diastereoselectivity, when THF as
solvent was employed (entry 7). This transformation proceeds
on a gram scale without noticeable loss of yield or selectivity.
With aldol product 2 in hand, further steps in the building
block synthesis were conducted (Scheme 2). Both acetylation
and levulination16 of the free hydroxyl group proceeded
(11) Collier, P. N.; Campbell, A. D.; Patel, I.; Raynham, T. M.; Taylor,
R. J. K. J. Org. Chem. 2002, 67, 1802.
(12) Evans, D. A.; Gage, J. R.; Leighton, J. L.; Kim, A. S. J. Org. Chem.
1992, 57, 1961.
(13) (a) Mahrwald, R. Modern Aldol Reactions: Wiley-VCH: Weinheim,
2004. (b) Ager, D. J.; Prakash, I.; Schaad, D. R. Aldrichim. Acta 1997, 30,
3.
(14) (a) Evans, D. A.; Bartroli, J.; Shih, T. L. J. Am. Chem. Soc. 1981,
103, 2127. (b) Jones, T. K.; Mills, S. G.; Reamer, R. A.; Askin, D.;
Desmond, R.; Volante, R. P.; Shinkai, I. J. Am. Chem. Soc. 1989, 111,
1157.
(9) Kawahara, K.; Sato, N.; Tsuge, K.; Seto, Y. Microbiol. Immunol.
2006, 50, 67.
(10) Ndonye, R. M.; Izmirian, D. P.; Dunn, M. F.; Yu, K. O. A.; Porcelli,
S. A.; Khurana, A.; Kronenberg, M.; Richardson, S. K.; Howell, A. R. J.
Org. Chem. 2005, 70, 10260.
(15) Crimmins, M. T.; King, B. W.; Tabet, E. A.; Chaudhary, K. J. Org.
Chem. 2001, 66, 894.
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Chem. Soc. 1975, 97, 1614. (b) van Boom, J. H.; Burgers, P. M. J.
Tetrahedron Lett. 1976, 17, 4875.
1574
Org. Lett., Vol. 10, No. 8, 2008

The synthesis of fatty acid H starts from commercially
available 1-tetradecene (Scheme 3). Treatment with AD-
Scheme 2. Synthesis of 5a, 5b, and 6
Scheme 3. Synthesis of Lipid F
smoothly to give the fully protected amides 3a and 3b in
quantitative yield. Treatment of thioacetal 3a with 1.8 equiv
of anisole and 5% trifluoroacetic acid in dichloromethane
resulted not only in the cleavage of the p-methoxybenzyl
ether, but also in concomitant cyclization. The desired
D-galacturonic acid thioglycoside 4a was afforded after this
transformation in 92% yield. Thioacetal 3b was converted
into 4b in 93% yield employing the same procedure. In
both cases, anomeric mixtures were separated by flash
column chromatography and the R-product was used in
further steps.
mix-R afforded diol 8 in 75% yield.22 The enantiomeric
excess23 of 78% is in line with observations by Sharpless et
al.22b for aliphatic alkenes with comparable chain length. Both
hydroxyl groups were differentiated by protection of the
primary alcohol with triphenylmethyl chloride. The second-
ary hydroxyl was then benzylated to furnish 9 in 75% yield,
over two steps. Trityl cleavage with TFA and triethylsilane
in methanol proceeded in 81% yield. Alcohol 10 was
subsequently oxidized using catalytic amounts of TEMPO
and bisacetoxyiodobenzene (BAIB) as cooxidant to give the
corresponding carboxylic acid H in 93% yield.24 Finally,
EDCI/HOBt-promoted amide coupling between amine G and
acid H yielded the desired diastereomer of ceramide F in
80% yield and at this stage the undesired diastereomer could
be separated.
With galacturonic acids 5a and 5b and ceramide F in hand,
glycosylation to yield conjugate 11 was studied (Table 2).
The main challenges, when performing these reactions, could
be attributed to the relatively poor solubility of ceramide F
in many solvent systems at low temperature. During these
studies, the well-known benefits of ether25 and the remote
anchimeric assistance26 of C4 esters in galacto-configured
systems in obtaining good R-selectivities became again
apparent, as omission led to a dramatic increase in â-gly-
coside formation. A compromise between yield and selectiv-
ity was found by employing acetyl-protected thioglycoside
Methanolysis of the chiral auxiliary in the presence of an
ester protecting group on C4 posed problems. Attempts to
conduct this transformation with sodium methoxide in
methanol17 or catalytic amounts of DMAP in methanol18 were
not successful. In both cases partial â-elimination of the C4
ester groups occurred. The reaction proceeded smoothly to
yield the methyl esters 5a and 5b in 80% and 81% yield,
respectively, when samarium(III) triflate in methanol (30 mol
%) was employed.19 No competing cleavage of the ester
groups was observed. Thus, both D-galacturonic acid thiogly-
cosides were obtained in only four steps from known8a and
readily accessible aldehyde E.
Thioethyl glycoside 5a was easily converted to the
alternative glycosylating agent, glycosyl phosphate 6, upon
NIS-promoted glycosidation with dibutyl phosphoric acid.
Deprotection of the C4 protecting groups to yield thiogly-
coside 7 proved to be facile. Both levulinoate and acetate
esters could be removed, with either hydrazine in acetic acid
and pyridine16b or in a methanolic HCl solution respectively.
Thus, this building block can be used for the assembly of
structures containing (1-4)-linked D-GalA.
(21) Characteristic coupling constants between H-4 and H-5 (J ) 1.3
Hz) as well as between H-3 and H-4 (J ) 3.4 Hz) unambiguously reveal
the desired galacto configuration of building block 5a. NOE correlations
between H-5 and H-4 as well as between H-4 and H-3 confirmed the
assignment. NOE spectra can be found in the Supporting Information.
(22) (a) He, L.; Byun, H. S.; Bittman, R. J. Org. Chem. 2000, 65, 7618.
(b) Kolb, H. C.; Vannieuwenhze, M. S.; Sharpless, K. B. Chem. ReV. 1994,
94, 2483.
At this stage, the diagnostic coupling constants20 served
to confirm the D-galacto configuration on R-thioglycoside
5a.21
(23) The % ee was determined by conversion of the diol to the bis ester
1
with (-)-R-Mosher’s reagent and subsequent H NMR analysis.
(17) Haigh, D.; Birrell, H. C.; Cantello, B. C. C.; Hindley, R. M.;
Ramaswamy, A.; Rami, H. K.; Stevens, N. C. Tetrahedron: Asymmetry
1999, 10, 1335.
(18) Baek, S.; Jo, H.; Kim, H.; Kim, S.; Kim, D. Org. Lett. 2005, 7, 75.
(19) (a) Evans, D. A.; Coleman, P. J.; Dias, L. C. Angew. Chem., Int.
Ed. Engl. 1997, 36, 2738. (b) Castagner, B.; Leighton, J. L. Tetrahedron
2007, 63, 5895.
(24) (a) Friestad, G. K.; Marie, J. C.; Deveau, A. M. Org. Lett. 2004, 6,
3249. (b) Kloth, K.; Bruenjes, M.; Kunst, E.; Joege, T.; Gallier, F.;
Adibekian, A.; Kirschning, A. AdV. Synth. Catal. 2005, 347, 1423.
(25) Wulff, G.; Ro¨hle, G. Angew. Chem., Int. Ed. Engl. 1974, 13, 157.
(26) (a) Miljkovic, M.; Yeagley, D.; Deslongchamps, P.; Dory, Y. L. J.
Org. Chem. 1997, 62, 7597. (b) Demchenko, A. V.; Rousson, E.; Boons,
G.-J. Tetrahedron Lett. 1999, 40, 6523. (c) Cheng, Y.-P.; Chen, H.-T.; Lin,
C.-C. Tetrahedron Lett. 2002, 43, 7721.
(20) Altona, C.; Haasnoot, C. A. G. Org. Magn. Reson. 1980, 13, 417.
Org. Lett., Vol. 10, No. 8, 2008
1575

Scheme 4. Global Deprotection of Glycosphingolipid A
Table 2. Glycosylation Studies
entry
donor
conditionsa
product
R/âb
yield (%)
1d
2d
3d
4d
5d
6d
7e
4a (R)
4a (R)
5b (R)
5b (R)
NIS,f TBSOTf g
NIS,f TfOHg
NIS,f TfOHg
DMTSTh
11c
11c
<10
45
49
<10
96
85
1.0:1
2.0:1
11b
11b
11ac
11ac
11ac
ester 13 in 74% yield. Finally, the acetate and methyl ester
of 13 were removed using lithium hydroperoxide in THF,
to yield fully deprotected glycosphingolipid A.
6 (R/â) TMSOTff
2.1:1
3.7:1
4.2:1
5a (R)
5a (R)
NIS,f TfOHg
NIS,f TfOHg
85
The chemistry presented here offers a convenient and high-
yielding route for the preparation of suitably protected
building blocks of D-galacturonic acid. Our strategy for the
de novo synthesis is based on the convergent connection of
linear synthetic intermediates, thus regioselective protection
steps as well as anomeric protection are no more necessary.
Consequently, orthogonally protected D-GalA thioglycoside
5a is obtained from known and easy available aldehyde E
in only four steps and 56% overall yield.
a Temperature -10 °C and 1.5 equiv of donor. b Ratio determined by
1H NMR. c The R- and â-products could be separated by column chroma-
tography. d Et2O-dichloroethane (1:1). e Dioxane-toluene (3:1). f 1.5 equiv.
g 0.15 equiv. h 6 equiv.
5a and the NIS/TfOH27 activator system. Thus, the product
11a was isolated in 85% yield and 4.2:1 selectivity with
dioxane/toluene (3:1) as solvent. Both anomers were sepa-
rated by flash column chromatography. To the best of our
knowledge, this result represents the most R-selective gly-
cosylation reaction between a galacturonic acid building
block and a ceramide reported so far.28
Having optimized the glycosylation reaction, all protecting
groups had to be removed. The initial plan was to cleave
first the acetate and the methyl ester of glycoconjugate 11a
in one step using lithium hydroperoxide in THF, followed
by removal of the benzyl groups with Pd(OH)2/C. The first
reaction proceeded in 93% yield, yet after hydrogenolysis
with either Pd/C or Pd(OH)2/C the desired alcohol A could
not be isolated (Scheme 4). The order of the two deprotection
steps was then inverted and debenzylation using Pearlman’s
catalyst in ethanol/chloroform was performed first to yield
These results should facilitate future syntheses of this
important class of immunological probes.
Application of the de novo strategy for rapid and efficient
preparation of other orthogonally protected carbohydrate
building blocks is currently under investigation and will be
reported in due course.
Acknowledgment. This research was supported by ETH
Zurich (TH Grant 16 06-3), a Fellowship from Studienstif-
tung des deutschen Volkes (to P.S.), and a Kekule´ Fellowship
from the Fonds der Chemischen Industrie (to A.A.).
Supporting Information Available: Experimental pro-
cedures and spectral characterization data for all new
compounds. This material is available free of charge via the
Internet at http://pubs.acs.org.
(27) Veebham, G. H.; van Leeuven, S. H.; van Boom, J. H. Tetrahedron
Lett. 1990, 31, 1331.
(28) For R-selective glycosylations with galacturonic acid, see: Nolting,
B.; Boye, H.; Vogel, C. J. Carbohydr. Chem. 2000, 19, 923.
OL800227B
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Org. Lett., Vol. 10, No. 8, 2008