Glycal glycosylation and 2-nitroglycal concatenation, a powerful combination for mucin core structure synthesis

Article abstract of DOI:10.1021/jo061670b

(Chemical Equation Presented) A 3,4-O-unprotected galactal derivative having bulky 6-O-TIPS protection (compound 2) could be regioselectively 3-O-glycosylated with O-(galactopyranosyl) trichloroacetimidates; depending on the protecting group pattern stere

Full text of DOI:10.1021/jo061670b

Glycal Glycosylation and 2-Nitroglycal Concatenation, a Powerful
Combination for Mucin Core Structure Synthesis
Ju¨rgen Geiger, B. Gopal Reddy, Gottfried A. Winterfeld, R. Weber, M. Przybylski, and
R. R. Schmidt*
Fachbereich Chemie, UniVersita¨t Konstanz, Fach M 725, D - 78457 Konstanz, Germany
Richard.Schmidt@uni-konstanz.de
ReceiVed August 11, 2006
A 3,4-O-unprotected galactal derivative having bulky 6-O-TIPS protection (compound 2) could be
regioselectively 3-O-glycosylated with O-(galactopyranosyl) trichloroacetimidates; depending on the
protecting group pattern stereoselectively R- and â-linked disaccharides were obtained. With O-(2-azido-
2-deoxyglucopyransyl) trichloroacetimidate as donor (compound 10A), glycosylation of 2 and of a 6-O-
unprotected galactal derivative led in acetonitrile as solvent exclusively to a â(1-3)- and a â(1-6)-
linked disaccharide, respectively. Nitration of the galactal moieties of the saccharides followed by Michael-
type addition of serine and threonine derivatives (7a,b) installed the R-galacto-configuration, thus readily
furnishing O-glycosyl amino acid building blocks for the incorporation of core 1, core 2, core 3, core 6,
and core 8 structures into glycopeptides. 2-Nitrogalactal and 2-nitroglucal derivatives could be also
successfully employed in glycoside bond formation via Michael-type addition in a reiterative manner,
affording the corresponding core 5, core 7, and core 6 building blocks. In this approach, highly
stereoselective glycoside bond formations were based exclusively on Michael-type addition to the nitro-
enol ether moiety of the 2-nitroglycals. Hence, 2-nitroglycals are versatile intermediates for base-catalyzed
glycoside bond formation.
Introduction
tosamine residue R-glycosidically linked to L-serine or L-threo-
nine (the TN-antigen).1,2 Eight core structures of mucin-type glyco-
peptides and the STN-antigen have been identified to date which
contain additional glycosyl residues at position 3 and/or position
6 to form complex O-glycans (Figure 1). These basic structures
are constituents of various glycoproteins, as for instance of the
antifreeze glycoproteins which are essential for the survival of
fish living under water freezing temperature conditions.3,4
The mucin glycoproteins have attracted much attention be-
cause of their fundamental importance in biological processes.1
The core structures bear at the reducing end an N-acetyl galac-
(1) (a) Varki, A. Glycobiology 1993, 3, 97-130. (b) Brockhausen, I.
Biochem. Biophys. Acta 1999, 1473, 67-95. (c) Brocke, C.; Kunz, H.
Bioorg. Med. Chem. 2002, 10, 3085-3112. (d) Devine, P. L.; McKenzie,
I. F. C. BioEssays 1992, 14, 619-625. (e) Gum, J. R., Jr. Biochem. Soc.
Trans. 1995, 23, 795-799.(f) Tsuboi, S.; Fukuda, M. BioEssays 2001, 23,
46-53. (g) Danishefsky, S. J.; Allen, J. R. Angew. Chem. 2000, 112, 882-
911; Angew. Chem., Int. Ed. 2000, 39, 836-863.
(2) Grogan, M. J.; Pratt, M. R.; Marcaurelle, L. A.; Bertozzi, C. R. Annu.
ReV. Biochem. 2002, 71, 593-634.
10.1021/jo061670b CCC: $37.00 © 2007 American Chemical Society
Published on Web 05/16/2007
J. Org. Chem. 2007, 72, 4367-4377
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Geiger et al.
SCHEME 1. Synthesis of Core 1 Building Blocks 9a,b
FIGURE 1. TN- and STN-antigens and the mucin core structures.
Previous approaches toward the construction of the charac-
teristic R-glycosidic linkage between 2-acetamido-2-deoxy-D-
galactopyranose and the side-chain hydroxy groups of L-serine
or L-threonine rely in most cases on the methodology introduced
by Paulsen in 19785 in which the nonparticipating azido group
is selected as latent amino function at position 2. This way
R-selective glycosylations with various donors and serine or
threonine acceptors could be successfully performed by various
groups.6 Also versatile 2-azido-2-deoxygalactose-based accep-
tors could be generated and employed for chain exten-
sion.6,7
Expanding the range of the previously reported efficient direct
glycosylation of particularly acid-sensitive galactal derivatives
with O-glycosyl trichloroacetimidates as glycosyl donors in the
presence of Sn(OTf)2 as mild catalyst8,9 and widening the scope
of the 2-nitro-glycal concatenation methodoloy10 should permit
highly versatile approaches toward the synthesis of all mucin
(3) (a) Harding, M. M.; Anderberg, P. I.; Haymet, A. D. J. Eur. J.
Biochem. 2003, 270, 1381-1392. (b) Yeh, Y.; Feeney, R. F. Chem. ReV.
1996, 96, 601-617. (c) Ben, R. N. ChemBioChem 2001, 2, 161-166.
(4) Tachibana, Y.; Fletcher, G. L.; Fujitani, N.; Monde, K.; Nishimura,
S.-I. Angew. Chem. 2004, 116, 874-880; Angew. Chem., Int. Ed. 2004,
43, 856-862.
core structures and derivatives thereof. The synthetic strategies
and the results are discussed in this paper.
(5) Paulsen, H.; Stenzel, W. Chem. Ber. 1978, 111, 2334-2347; Paulsen,
H.; Stenzel, W. Chem. Ber. 1978, 111, 2348-2357; Paulsen, H.; Kolar,
C.; Stenzel, W. Chem. Ber. 1978, 111, 2358-2369; Paulsen, H.; Kolar, C.
Chem. Ber. 1979, 112, 3190-3202.
Results and Discussion
(6) (a) Ferrari, B.; Pavia, A. A. Carbohydr. Res. 1980, 79, C1-C7. (b)
Paulsen, H.; Ho¨lck, J.-P. Carbohydr. Res. 1982, 109, 89-107.(c) Grundler,
G.; Schmidt, R. R. Liebigs Ann. 1984, 1826-1847. (d) Kinzy, W.; Schmidt,
R. R. Carbohydr. Res. 1987, 164, 265-276. (e) Paulsen, H.; Rauwald, W.;
Weichert, U. Liebigs Ann. 1988, 75-86. (f) Kinzy, W.; Schmidt, R. R.
Carbohydr. Res. 1989, 193, 33-47. (g) Nakahara, Y.; Iijima, H.; Sibayama,
S.; Ogawa, T. Tetrahedron Lett. 1990, 31, 6897-6900. (h) Rademann, J.;
Schmidt, R. R. Carbohydr. Res. 1995, 269, 217-225. (i) Chen, ×.-T.;
Sames, D.; Danishefsky, S. J. J. Am. Chem. Soc. 1998, 120, 7760-7769.
(j) Tsudu, T.; Nishimura, S. I. Chem. Commun. 1996, 2779-2780. (k)
Schwarz, J. K.; Kuduk, S. D.; Chen, ×.-T.; Sames, D.; Glunz, P. W.;
Danishefsky, S. J. J. Am. Chem. Soc. 1999, 121, 2662-2673. (l) Marcurelle,
L. T.; Bertozzi, C. R. J. Am. Chem. Soc. 2001, 123, 1487-1505. (m) Tseng,
P. H.; Jiang, W. T.; Chang, M. Y.; Chen, S. T. Chem. Eur. J. 2001, 7,
498-510.
Galactal Glycosylation followed by 2-Nitrogalactal Con-
catenation. The mucin core structure synthesis requires R- and
â(1-3)-glycosidic linkages of galactose (Gal) to N-acetylga-
lactosamine (GalNAc) (core 1 and 8), â(1-3)-and â(1-6)-
linkages of N-acetyl glucosamine (GlcNAc) to GalNAc (core 3
and 6), R(1-3)- and R(1-6)-linkages of GalNAc to GalNAc
(core 5 and 7), and formation of branched trisaccharides having
â(1-3)-linkage of Gal or GlcNAc and â(1-6)-linkages of
GlcNAc to GalNAc (core 2 and 4) (Scheme 1). Hence, for the
galactal glycosylation-nitrogalactal concatenation strategy gen-
erating two glycosidic linkages, as shown in Figure 2, first R-
and â-selective glycosylation of galactal at 3-O and/or 6-O under
mild conditions is required (step 1). As reported,8 this demand
is already fulfilled for the 3-O-galactosylation of galactal with
the help of a standard glycosyl donor (step 1a). However, for
(7) Herzner, H.; Reipen, T.; Schulz, M.; Kunz, H. Chem. ReV. 2000,
100, 4495-4538.
(8) Geiger, J.; Barroca, N.; Schmidt, R. R. Synlett 2004, 836-840.
(9) Glycosylation methods using glycals as acceptors and as donors
employing 3,3-dimethyldioxirane for the activation of the double bond has
been developed. See for example: Halcomb, R. L.; Danishefsky, S. J. J.
Am. Chem. Soc. 1989, 111, 6661-6666; Liu, K. K.-C.; Danishefsky, S. J.
J. Org. Chem. 1994, 59, 1892-1894; Zheng, C.; Seeberger, P. H.;
Danishefsky, S. J. J. Org. Chem. 1998, 63, 1126-1130; Danishefsky, S.
J.; Bilodeau, M. T. Angew. Chem. 1996, 108, 1482-1522; Angew. Chem.,
Int. Ed. 1996, 35, 1380-1419.
(10) (a) Winterfeld, G. A.; Schmidt, R. R. Angew. Chem. 2001, 113,
2718-2721; Angew. Chem. Int. Ed. Engl. 2001, 40, 2654-2657. (b)
Winterfeld, G. A.; Khodair, A. I.; Schmidt, R. R. Eur. J. Org. Chem. 2003,
1009-1021.
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Mucin Core Structure Synthesis
SCHEME 2. Synthesis of Core 2 Building Block 14
FIGURE 2. Generation of two glycosidic linkages via galactal
glycosylation followed by 2-nitrogalactal concatenation.
the introduction of the GalNAc residue (core 5 and 7) and
particularly for the introduction of the less reactive GlcNAc
residue (leading to core structures 2, 3, 4, and 6), suitable
glycosyl donors and reaction conditions have to be developed.
The regioselectivity of 3-O and/or 6-O-glycosylation of galactal
can be controlled with minimal protecting group manipulations
because these two hydroxy groups can be readily differentiated
in this acceptor.11 Second, nitro group introduction at C-2 of
the galactal moiety has to be performed via a highly selective
electrophilic substitution reaction (step 2). Third, Michael-type
addition of alcohols and particularly of serine (Ser) and threonine
(Thr) derivatives as nucleophiles at the anomeric carbon of the
nitrogalactal moiety should be R-selective with the nitro group
at C-2 adopting after protonation the equatorial position. Thus,
it was perceived that via substrate stereocontrol two stereogenic
centers will be generated, hopefully leading directly to the
desired R-galacto-configured compounds (step 3). Fourth,
transformation of the nitro group into an amino group will
provide the target compounds (step 4). The application of this
synthesis strategy (via steps 1a, 2, 3, and 4 in this part) is
demonstrated in successful synthesis of core 1 and core 8 mucin
structures where it is particularly suitable. However, core 2, core
3, and core 6 building blocks could also be efficiently obtained
using this strategy.
desired adducts 8a,b in 65% yield each (1H NMR: J1a,2a ) 3.6
Hz). Reduction of the nitro group with zinc/HCl8,14 in a water-
acetic acid-THF mixture at 0 °C and then N-acetylation with
acetic anhydride in pyridine furnished target molecules 9a,b in
only five steps.
Disaccharide 5, after 6a-O-desilylation with tetrabutylam-
monium fluoride (TBAF) in THF, furnished 11, which is
obviously also an ideal precursor for the synthesis of core 2
building blocks, if 6a-O-glycosylation of the galactal moiety
of 11 with a glucosamine precursor can be performed. To this
end, known O-(2-azido-2-deoxy-glucopyranosyl) trichloroace-
timidate 10A15 was selected as reactive glycosyl donor which
in acetonitrile at low temperature was expected to lead to
â-linkage due to the nitrile effect (Scheme 2).16 As expected,
under these conditions activation of 10A with tin(II) triflate was
still possible, and this way the desired â-linked trisaccharide
12 was obtained exclusively in good yield (1H NMR: J1c,2c
)
As designed, glycosylation of 3,4-O-unprotected galactal 212
with galactosyl donor 113 in the presence of Sn(OTf)2 as catalyst
in CH2Cl2 at 0 °C afforded the â-linked disaccharide 3 in 75%
yield (1H NMR: J1b,2b ) 8.3 Hz). Direct O-benzylation of 3
led to 4-O-benzyl derivative 4. However, nitration of this
compound having a 2b-O-acetyl group was met with difficulties
(Scheme 1). Removal of the 2b-O-acetyl group from 3 followed
by O-benzylation furnished 5 which on nitration via addition-
elimination at the enol ether moiety led cleanly to 6 having the
nitrogalactal moiety for Michael-type addition of commercially
available serine and threonine derivatives 7a,b. Treatment of 6
with 7a or 7b in the presence of potassium tert-butoxide in
toluene proved to be most efficient in shifting the Michael
addition-elimination equilibrium to the product side. Thus, the
R-galacto-configuration was exclusively installed, affording the
br s). Nitration of 12 (to form 13) and serine 7a addition under
standard conditions afforded target molecule 14a with R-linked
serine as the only product (1H NMR: J1a,2a ) 4.6 Hz). The
ease of azido and nitro group transformation into N-acetylamino
groups will be demonstrated for another example (see below).
An efficient method for the R-selective 3-O-galactosylation
of 6-O-triisopropylsilyl (TIPS)-protected galactal 2 as acceptor
with O-benzyl-protected galactosyl donor 1517 is not yet
available.8 However, variations of solvent and temperature
conditions exhibited that the R/â-ratio can be greatly improved
by performing the reaction in a dichloromethane/ether mixture18
with tin(II) triflate as catalyst at room temperature leading to a
(14) (a) Brown, H. C.; Brown, C. A. J. Am. Chem. Soc. 1963, 85, 1005-
1006. (b) Geiger, J. Dissertation, Universita¨t Konstanz, 2005.
(15) Kinzy, W.; Schmidt, R. R. Liebigs Ann. Chem. 1985, 1537-1545.
(16) Schmidt, R. R.; Behrendt, M.; Toepfer, A. Synlett 1990, 694-697.
(17) Wegmann, B.; Schmidt, R. R. J. Carbohydr. Chem. 1987, 357-
375.
(11) Kinzy, W.; Schmidt, R. R. Tetrahedron Lett. 1987, 28, 1981-1984.
(12) Gervay, J.; Peterson, J. M.; Oriyama, T.; Danishefsky, S. J. J. Org.
Chem. 1993, 58, 5465-5468.
(13) Sakai, K.; Nakahara, Y.; Ogawa, T. Tetrahedron Lett. 1990, 31,
3035-3038.
(18) Schmidt, R. R. Angew. Chem., Int. Ed. 1998, 25, 212-235; and
references therein.
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Geiger et al.
SCHEME 4. Synthesis of Core 3 Building Block 28a
SCHEME 3. Synthesis of Core 8 Building Blocks 22 and
24a,b
triflate as catalyst at -40 °C exclusively the desired â-linked
disaccharide 25A could be obtained in good yield (1H NMR:
J1b,2b ) 8.6 Hz). Even glucosamine donors 10B,19 10C,20
requiring generally stronger activation conditions than 10A,
reacted smoothly with acceptor 2 in dichloromethane as solvent
at 0 °C to afford disaccharides 25B,C, respectively, in similar
yield as 25A. 4a-O-Benzylation of 25A (f 26) and then
nitration led to C-2a-nitro derivative 27. Addition of serine
derivative 7a in the presence of potassium tert-butoxide in
toluene gave exclusively R-galacto-configured core 3 building
block 28a (1H NMR: J1a,2a ) 3.8 Hz). Similar to the core 2
building block synthesis (Scheme 2) galactal containing disac-
charide 26 is also an ideal precursor for core 4 building block
synthesis.
With these results in hand, it was perceived that the synthesis
of core 6 building blocks should be also readily achievable. To
this end, known galactal derivative 2921 was 6-O-desilylated
with TBAF in THF furnishing the required 6-O-unprotected
acceptor 30 (Scheme 5) which on reaction with donor 10A in
acetonitrile at -40 °C under tin(II) triflate catalysis led only to
â-linked disaccharide 31 in good yield (1H NMR: J1b,2b ) 7.2
Hz). Nitration of 31 to 32 and then base-catalyzed addition of
7a,b afforded exclusively the desired R-linked disaccharides
33a,b in 64% yield each. For these examples, also azido and
nitro group reduction was investigated. Treatment of 33a,b with
thioacetic acid led cleanly to azide transformation into acety-
lamino groups furnishing compounds 34a,b. Nitro group reduc-
tion in 34a,b could be performed by treatment with platinized
Raney-Ni T422 under hydrogen pressure followed by treatment
10:1 16R/16â ratio in 52% yield (1H NMR, 16R: J1b,2b ) 3.8
Hz; 16â: 1H NMR: J1b,2b ) 7.8 Hz) (Scheme 3). 4a-O-
Benzylation of 16R under standard conditions to form 17
followed by nitration led to the disaccharide 19, which contains
the 2-nitrogalactal moiety. 6a-O-Desilylation of 16R and then
4a,6a-O-dibenzylation afforded per-O-benzylated disaccharide
18, which on nitration provided the desired C-2a-nitro derivative
20. Both compounds, 19 and 20, were subjected to base-
catalyzed addition of serine and threonine derivatives 7a,b. Thus,
20 was readily reacted with 7a to give the desired R-galacto-
configured addition product 21a (1H NMR: J1a,2a ) 3.8 Hz)
indicating that the high stereocontrol is not only supported by
sterically demanding 6a-O-groups but also by corresponding
3a-O-groups. Similarly, from 19 and 7a,b R-galacto-configured
products 23a,b were obtained (1H NMR: J1a,2a ) 4.2 Hz). Nitro
group reduction of 21a and 23a,b with Zn/HCl in a water/acetic
acid/THF mixture afforded the core 8 building blocks 22a and
24a,b, respectively.
(19) Dullenkopf, W.; Castro-Palomino, J. C.; Manzoni, L.; Schmidt, R.
R. Carbohydr. Res. 1996, 296, 135-147.
Further extension of this method to â-selective attachment
of glucosamine donor 10A to 6-O-protected galactal 2 as
acceptor would permit the synthesis of core 3 building blocks
(Scheme 4). Indeed, with acetonitrile as solvent and tin(II)
(20) Castro-Palomino, J. C.; Schmidt, R. R. Tetrahedron Lett. 1995, 36,
5343-5346.
(21) Timmers, C. M.; Wigchert, S. C. M.; Leeuwenburgh, M. A.; Marel,
G. A. v. d.; van Boom, J. H. Eur. J. Org. Chem. 1998, 91-92.
4370 J. Org. Chem., Vol. 72, No. 12, 2007

Mucin Core Structure Synthesis
SCHEME 5. Synthesis of Core 6 Building Blocks 35a,b
SCHEME 6. Synthesis of Core 7 Building Blocks 41a and
42a
of the intermediates with acetic anhydride pyridine afforded the
target compounds 35a,b in very good yields.
Reiterative 2-Nitroglycal Concatenation. The presence of
GalNAcR(1-3)- and GalNAcR(1-6)GalNAc linkages in mucin
core 5 and core 7, respectively, provides an interesting alterna-
tive to the above-reported glycoside bond formation methodol-
ogy: glycosidations can be exclusively based on base-catalyzed
2-nitroglycal concatenation (Figure 2, step 1b). This strategy
(via steps 1b, 2, 3, and 4 in this part) should be particularly
suitable for core 5 and core 7 syntheses if not only primary
sugar hydroxy groups (as required for core 7) but also secondary
sugar hydroxy groups (as in core 5) can be added to the
2-nitroglycal moiety. This way, synthesis strategies starting both
from the reducing end or from the nonreducing end can be
undertaken. This synthetic design should also be useful, for
instance, in core 3 and core 6 syntheses if 2-nitroglucal can be
successfully integrated in this 2-nitroglycal concatenation
scheme.
The synthesis of a core 7 building block (see Scheme 6) starts
from the reducing end. Known galactal derivative 3621 was
transformed under standard conditions into C-2-nitro derivative
37. Addition of serine and threonine derivatives 7a,b in the
presence of potassium tert-butoxide as base in toluene afforded
exclusively the desired 2-nitro-2-deoxy-R-d-galactopyranosides
38a,b (1H NMR: J1,2 ) 4.2 Hz). Cleavage of the 6-O-tert-
butyl-diphenylsilyl (TBDPS) group with TBAF in the presence
of acetic acid in THF furnished 6-O-unprotected derivatives
39a,b. However, investigations toward addition of 39a to 37 in
the presence of potassium tert-butoxide in toluene led to
â-elimination of 7a and its readdition to 37 furnishing 38a;
practically no disaccharide was obtained. In order to avoid this
reaction, the nitro group in 39a was transformed into an
N-acetylamino group as described above, affording the TN-
antigen building block 40a. Investigation of Michael-type
addition of this compound to 37 led cleanly to the desired
disaccharide 41a having the required R-galacto configuration
(1H NMR: J1b,2b ) 4.3 Hz). Hydrogenation of the nitro group
with platinized Raney-Ni T4 in ethanol, then N-acetylation,
hydrogenolytic cleavage of the O-benzyl groups and per-O-
acetylation afforded after four steps the core 7 building block
42a in 56% yield.
The decisive step in core 5 building block synthesis, based
on reiterative nitrogalactal concatenation starting from the
nonreducing end, is successful Michael-type addition of second-
ary sugar hydroxy groups to 2-nitrogalactal derivatives. To this
end, base-catalyzed reaction of 37 with 3,4-O-unprotected
galactal 2 was investigated. To our delight, the desired R(1-
3)-linked disaccharide 43 was cleanly obtained under standard
conditions (1H NMR: J1b,2b ) 4.2 Hz) (Scheme 7). However,
3,4,6-tri-O-benzyl-2-nitrogalactal afforded with 2 a 3:1-mixture
of the R/â-anomers.14b Thus, it appears that the bulky 6-O-
TBDPS group in 37 supported R-addition which is also favored
for stereoelectronic reasons. The 4a-hydroxy group in 43 was
acetylated to obtain 44 whose nitration proceeded well in the
presence of the axial acetoxy group affording 2-nitro derivative
45 in 86% yield. Also Michael-type addition of 7a could be
carried out under standard conditions in the presence of the 4a-
O-acetyl group furnishing the desired R-anomer 46a (1H
NMR: J1a,2a ) 4.2 Hz) in 74% yield. One-pot transformation
of the two nitro groups into N-acetylamino groups could be
carried out on hydrogenation with platinized Raney-Ni T4 as
catalyst and then N-acetylation with acetic anhydride in pyridine
(22) Nishimura, S. Bull. Chem. Soc. Jpn. 1959, 32, 61-64.
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Geiger et al.
SCHEME 7. Synthesis of Core 5 Building Block 47a
SCHEME 8. Synthesis of Core 6 Building Block 55a
FIGURE 3. Conformation of 51.
addition to 2-nitroglucal derivatives as well, for which in earlier
studies even with simple alcohols as nucleophiles mainly modest
results were obtained.26 To this end, a 2-nitroglucal derivative
51 was designed having 3,6-di-O-benzyl and sterically demand-
ing 4-O-tert-butyldimethylsilyl (TBS) protection on the R-face
(Scheme 8). For the synthesis of compound 51, glucal 4811 was
converted to 4927 via O-desilylation, O-benzylation, and acid-
catalyzed tetrahydropyranyl (THP) cleavage. Introduction of the
4-O-TBS group to get 50 followed by nitration furnished 51. It
was expected that the bulky TBS group together with the allylic
4
effect would favor transition of the H5 conformer of 51 into
the 5H4 conformer with essentially axial substituents at C-3, C-4,
and C-5. This way, as shown in Figure 3 for steric and
stereoelectronic reasons nucleophilic attack at C-1 should
preferably take place from the â-side. This assumption was
1
supported by the H NMR data of 51 (J3,4 ) 2.2 Hz) and by
affording core 5 building block 47a. Reduction of 46a with SmI2
in a mixture of methanol-THF23 afforded the 2a,2b-dihy-
droxyamino derivative which on treatment with pyruvic acid
in aqueous DMF24 furnished also 47a in better yield.
So far, the 2-nitroglycal concatenation was essentially based
on 2-nitrogalactal derivatives, which, in the presence of a
suitable protecting group pattern, led with alcohols as nucleo-
philes cleanly to R-galacto-configured adducts. However, under
variation of the base and/or the nucleophile also â-galacto-
configured adducts could be obtained preferentially or even
exclusively.25 Extension of the reiterative nitroglycal concatena-
tion to other mucin core structures would require â-selective
the following reaction: Base-catalyzed addition of 6a-O-
unprotected galactal 52, having also bulky 3-O-and 4-O-silyl
protecting groups, led exclusively to â-gluco-configured product
53 in which the b-ring adopted the 4C1 conformation (1H
NMR: J1b,2b ) 8.1, J3,4 ) 8.9, J4,5 ) 8.9 Hz). Nitration of 53
under standard conditions forming 54 and then Michael-type
addition of 7a afforded the desired core 6 building block 55a
in good yield (1H NMR: J1a,2a ) 4.1 Hz). Hence, extension of
(25) Das, J.; Schmidt, R. R. Eur. J. Org. Chem. 1998, 1609-1613. (b)
Winterfeld, G. A.; Das, J.; Schmidt, R. R. Eur. J. Org. Chem. 2000, 3047-
3050. (c) Barroca, N.; Schmidt, R. R. Org. Lett. 2004, 6, 1551-1554.
(26) (a) Lemieux, R. U.; Nagabushan, T. L.; O’Neill, I. K. Can. J. Chem.
1968, 46, 413-418. (b) Takamoto, T.; Sudoh, R.; Nakagawa, T. Carbohydr.
Res. 1973, 27, 135-140.
(23) Kende, A. S.; Mendoza, J. S. Tetrahedron Lett. 1991, 32, 1699-
1702.
(24) (a) Chow, K. Y.-S.; Bode, J. W. J. Am. Chem. Soc. 2004, 126,
8126-8127. (b) Sohn, S. S.; Bode, J. W. Org. Lett. 2005, 7, 3873-3876.
(27) Danishefsky, S. J.; Behar, V.; Randolph, J. T.; Lloyd, K. O. J. Am.
Chem. Soc. 1995, 117, 5701-5711.
4372 J. Org. Chem., Vol. 72, No. 12, 2007

Mucin Core Structure Synthesis
mol) and selectively protected galactal 212 (3.37 g, 0.012 mol) were
dissolved in CH2Cl2 (60 mL) and cooled to 0 °C. Then a mixture
of tin(II) triflate (140 mg, 0.36 mmol) and MeCN (3 mL) was added
dropwise within 15 min. After 30 to 60 min (TLC-monitoring),
the reaction was stopped by adding triethylamine. The organic phase
was removed under reduced pressure. Purification of the residue
by flash chromatography (petroleum ether/ethyl acetate ) 6:1)
afforded disaccharide 3 (8.2 g, 0.011 mol, 75%) as a slightly yellow
oil. Rf ) 0.44 (petroleum ether/ethyl acetate ) 3:1); [R]D ) -5.1
this methodology to 2-nitroglucals could be successfully
demonstrated.
Conclusion
The 2-nitroglycal concatenation concept for glycoside bond
formation consisting of mild acid-catalyzed O-glycosylation of
glycals, nitration of the enol ether moiety to generate Michael-
type acceptors, highly stereoselective addition of nucleophiles,
particularly of low reactive alcohols, and then nitro group
reduction to the amino group could be successfully applied to
the synthesis of practically all mucin core structures. This way,
R- and â-selective glycosylation of galactals with galactopyra-
nosyl and 2-amino-2-deoxy-glucopyranosyl donors could be
obtained. The concept is also accessible to reiteration, thus
permitting the linkage of GlcNAc and GalNAc residues in
various combinations. The high stereocontrol in the Michael-
type addition is based on stereoelectronic and/or steric effects.
Hence, the selection of the protecting group pattern plays an
important role.
Glycosidically linked galactose, glucose, and 2-amino-2-
deoxy-galactose and -glucose residues are basic constituents of
practically all mammalian oligosaccharides and not only of the
mucin core structures. Hence, the base-catalyzed nitroglycal
concatenation concept has a very wide scope, and it is a valuable
addition to the many acid-promoted methodologies developed
for glycoside bond formation.18,28
1
(c ) 1, CHCl3). H NMR (600 MHz, CDCl3): δ ) 7.37-7./(m,
15 H, arom. H), 6,37 (d, J1,2 ) 6.2 Hz, 1 H, 1a-H), 5.39 (dd, J2,1
) 8.3 Hz, J2,3 ) 10.1 Hz, 1 H, 2b-H), 4.95.(d, J ) 11.6., 1 H,
benzyl H), 4,69 (d, J ) 12.2 Hz, 1 H, benzyl H), 4.62-4.58 (m, 2
H, 2a-H, benzyl H), 4.54-4.51 (m, 2 H, 1b-H, benzyl H), 4.46-
4.44 (m, 2 H, benzyl H), 4.38 (s, 1 H, 3a-H), 4.14 (s, 1 H, 4a-H),
4.03-3.99 (m, 2 H, 6a-H, 4b-H), 3.90-3.87 (m, 2 H, 5a-H, 6a′-
H), 3.64-3.55 (m, 4 H, 3b-H, 5b-H, 6b-H, 6b′-H), 2.76 (s, 1 H,
OH), 2.04 (s, 3 H, OAc), 1.13-1.03 (m, 21 H, TIPS). 13C NMR
(151 MHz, CDCl3): δ ) 169.4 (C-Ac), 145.0 (C-1a), 138.2-127.2
(18 C, arom. C), 100.5 (C-1b), 98.7 (C-2a), 80.5, 80.0 (C-3b), 76.9
(C-5a), 75.0, 74.5, 73.7 (C-5b), 73.0, 72.7 (C-3a), 72.4 (C-4b), 71.3
(C-2b), 68.2 (C-6b), 63.3 (C-4a), 61.6 (C-6a), 20.9-11.8 (9 C).
MALDI-MS (positive mode, DHB): [M + Na]+: m/z calcd.:
800.0, found: 800.0. C44H60SiO10 (776.98) calcd.: C: 68.02, H:
7.72, found: C: 67.67, H: 7.86.
O-(2-O-Acetyl-3,4,6-tri-O-benzyl-â-D-galactopyranosyl)-(1f3)-
3-O-benzyl-6-O-(triisopropylsilyl)-D-galactal (4). To a solution
of 3 (1.0 g, 1.29 mmol) in DMF (5 mL) was added NaH (53 mg,
2.21 mmol). After 15 min of stirring, BnBr (0.17 mL, 1.42 mmol)
was added dropwise. After termination of the reaction, ethyl acetate
was added (10 mL) and the organic phase washed (3 × 5 mL H2O),
dried with MgSO4, and concentrated under reduced pressure.
Purification by flash chromatography (petroleum ether/ethyl acetate
) 7.5:1) afforded entirely protected disaccharide 4 (624 mg, 0.72
mmol, 56%) as a slightly yellow oil. Rf ) 0.80 (petroleum ether/
ethyl acetate ) 2:1); [R]D ) -29.7 (c ) 1, CHCl3). 1H NMR (250
Experimental Section
General Procedure for the Nitration of Glycals (A). Concen-
trated nitric acid (5 mL, 79 mmol) was added dropwise to acetic
anhydride (50 mL) at 10 °C (temperature of the solution) under
constant stirring. Once the addition was complete, the solution was
cooled to -30 °C. Then a solution of the glycal (20.7 mmol) in
acetic anhydride (10 mL) was added over a period of 10-15 min,
and the mixture was stirred at this temperature for 1 h. The reaction
mixture was poured onto ice-water (150 mL). Brine (100 mL)
was added, and the aqueous layer was extracted with diethyl ether
(3 × 150 mL). The combined organic extracts were dried over
sodium sulfate and the solvents removed by coevaporation with
toluene. The crude 2-nitro sugar intermediate was dissolved in
dichloromethane (10 mL) and slowly added to an ice-cold, stirred
solution of triethylamine (3.5 mL, 25 mmol) in dichloromethane
(10 mL). After complete addition, the cooling bath was removed
and stirring continued for 25 min. The organic phase was washed
with 1 n HCl solution and dried over sodium sulfate. Removal of
the solvent in vacuo and then silica gel chromatography of the crude
furnished 2-nitroglycals.
General Procedure for the Transformation of the Nitro
Group to the Acetamido Group (B). The nitro compound (1.0
mmol) was dissolved in a mixture of THF (75 mL), concd HCl (3
mL), HOAc (18 mL), and H2O (30 mL) and cooled to 0 °C. Zinc
dust (24 mmol) was added. After stirring for 2 h at 0 °C, excess
zinc(II) dust was removed by filtration; the reaction mixture was
diluted with CH2Cl2, washed with H2O, sat. aq NaHCO3, and H2O,
and dried over anhyd MgSO4. Evaporation of the solvents and
acetylation of the residue with acetic anhydride/pyridine (2:1)
followed by concentration of the mixture and purification by silica
gel chromatography afforded the N-acetamido glycoside.
MHz, CDCl3): δ ) 7.40-7.22 (m, 20 H, arom. H), 6.33 (d, J1,2
)
6.4 Hz, 1 H, 1a-H), 5.48 (dd, J2,1 ) J2,3 ) 8.0 Hz, 1 H, 2b-H),
4.98 (d, J ) 11.0 Hz, 1 H, benzyl H), 4.97 (d, J ) 11.0 Hz, 1 H,
benzyl H), 4.72-4.51 (m, 7 H, 1b-H, 2a-H, 3a-H, benzyl H), 4.45
(s, 2 H, benzyl H), 4.01 (br s, 2 H, 4a-H, 5a-H), 3.92-3.92 (m, 1
H, 4b-H), 3.68 (d, 2 H, 6a-H, 6a′-H), 3.63-3.54 (m, 4 H, 3b-H,
5b-H, 6b-H, 6b′-H), 1.99 (s, 3 H, OAc), 1.05 (m, 21 H, TIPS).
MALDI-MS (positive mode, DHB): [M + Na]+: m/z calcd.:
890.1, found: 890.4. C51H66O10Si (867.11) calcd.: C: 70.64, H:
7.67, found: C: 70.64, H: 7.57.
O-(2,3,4,6-Tetra-O-benzyl-â-D-galactopyranosyl)-(1f3)-3-O-
benzyl-6-O-(triisopropylsilyl)-D-galactal (5). 3 (1.6 g, 2.06 mmol)
was dissolved in 30 mL methanol and added a spatula sodium
methanolate. After termination of the reaction, the solvent was
evaporated in vacuo. The residue was then dissolved in DMF, and
NaH (236 mg, 10.0 mmol) and BnBr (0.6 mL, 4.53 mmol) were
added. The reaction was terminated after 16 h by adding ethyl
acetate. The organic phase was washed with water, then dried with
MgSO4, and finally the solvents removed under reduced pressure.
Purification by flash chromatography (toluene/ethyl acetate ) 10:
1) furnished compound 5 (1.4 g, 1.60 mmol, 78%) as slightly yellow
oil. Rf ) 0.79 (petroleum ether/ethyl acetate ) 4:1); [R]D ) +28.0
(c ) 1, CHCl3). 1H NMR (250 MHz, CDCl3): δ ) 7.30-7.22 (m,
25 H, arom. H), 6.23 (dd, J1,2 ) 6.2 Hz, J1,3 < 1.0 Hz, 1 H, 1a-H),
5.13 (d, J ) 11.8 Hz, 1 H, benzyl H), 5.01 (d, J ) 11.2 Hz, 1 H,
benzyl H), 4.97 (d, J ) 11.2 Hz, 1 H, benzyl H), 4.82-4.62 (m, 7
H, 2a-H, 3a-H, benzyl H), 4.59 (d, J1,2 ) 7.6 Hz, 1 H, 1b-H), 4.44
(s, 2 H, benzyl H), 4.13 (br s, 1 H, 5a-H), 3.97-3.83 (m, 4 H,
4a-H, 4b-H, 2b-H, 6-H), 3.68-3.55 (m, 5 H, 3 × 6-H, 5b-H, 3b-
H), 1.05-1.02 (m, 21 H, TIPS). MALDI-MS (positive mode,
DHB): [M + Na]+: m/z calcd.: 938.3, found: 939.4. C56H70O9Si
(915.27 × H2O) calcd.: C: 72.07, H: 7.78, found: C: 72.00, H:
7.83.
O-(2-O-Acetyl-3,4,6-tri-O-benzyl-â-D-galyctopyranosyl)-(1f3)-
6-O-(triisopropylsilyl)-D-galactal (3). Imidate 113 (9.0 g, 0.014
(28) (a) Schmidt, R. R.; Kinzy, W. AdV. Carbohydr. Chem. Biochem.
1994, 50, 21-123. (b) Toshima, K.; Tatsuta, K. Chem. ReV. 1993, 93,
1503-1531. (c) Davis, B. G. J. Chem. Soc., Perkin Trans. 1 2000, 2137-
2160. (d) Seeberger, P. H.; Hase, W. Chem. ReV. 2000, 100, 4349-4394.
J. Org. Chem, Vol. 72, No. 12, 2007 4373

Geiger et al.
O-(2,3,4,6-Tetra-O-benzyl-â-D-galactopyranosyl)-(1f3)-3-O-
benzyl-2-nitro-6-O-(triisopropylsilyl)-D-galactal (6). Following
general procedure A from 5 (320 mg, 0.35 mmol), after flash
chromatography with toluene/ethyl acetate (40:1) compound 6 (228
mg, 0.24 mmol, 68%) was obtained as a slightly yellow oil. Rf )
0.50 (toluene/ethyl acetate ) 5:1); [R]D ) +3.5 (c ) 0.25, CHCl3).
1H NMR (250 MHz, CDCl3): δ ) 8.08 (s, 1 H, 1a-H), 7.29-7.21
(m, 25 H, arom. H), 5.09 (d, J3,4 ) 4.6 Hz, 1 H, 3a-H), 5.05-5.00
(m, 2 H, benzyl H), 4.95 (d, J1,2 ) 7.7 Hz, 1 H, 1b-H), 4.74-4.52
(m, 6 H, benzyl H), 4.34 (s, 2 H, benzyl H), 4.15-4.08 (m, 1 H,
5a-H), 3.99 (dd, J4,3 ) 4.6 Hz, J4,5 ) 2.7 Hz, 1 H, 4a-H), 3.89 (d,
J ) 2.6 Hz, 1 H, 4b-H), 3.77 (dd, J2,1 ) 7.7, J2,3 ) 10.4 Hz,1 H,
2b-H), 3-67-3.50 (m, 6 H, 3b-H, 5b-H, 6a-H, 6a′-H, 6b-H, 6b′-
H), 0.98-0.97 (m, 21 H, TIPS). MALDI-MS (positive mode,
DHB): [M + Na]+: m/z calcd.: 982.1, found: 982.4. C56H69NO11-
Si (960.23) calcd.: C: 70.05, H: 7.24, N: 1.46, found: C: 70.44,
H: 7.17, N: 1.53.
O-(2,3,4,6-Tetra-O-benzyl-â-D-galactopyranosyl)-(1f3)-(4-O-
benzyl-2-deoxy-2-nitro-6-(triisopropylsilyl)-r-D-galactopyrano-
syl)-N-(tert-butyloxycarbonyl)-L-serine tert-Butyl Ester (8a). 6
(111 mg, 0.12 mmol) and 7a (70 mg, 0.25 mmol) were dissolved
in 5 mL of dry toluene. Then potassium tert-butoxide (3 mg, 0.026
mmol) was added under stirring. After 2 h the reaction was
neutralized with acetic acid and the solvent removed in vacuo.
Purification by flash chromatography with toluene/ethyl acetate (40:
1) furnished compound 8a (95 mg, 0.08 mmol, 65%) as a colorless
oil. Rf ) 0.53 (petroleum ether/ethyl acetate ) 5:1); [R]D ) +17.9
(c ) 1, CHCl3) ) +17.9. 1H NMR (600 MHz, CDCl3): δ ) 7.32-
7.16 (m, 25 H, arom. H), 5.27 (d, J1,2 ) 3.6 Hz, 1 H, 1a-H), 5.18
(d, J ) 8.4 Hz, 1 H, Ser-NH), 5.00 (d, J ) 12.0 Hz, 1 H, benzyl
H), 4.93 (dd, J2,1 ) 3.6, J2,3 ) 10.8, 1 H, 2a-H), 4.90 (d, J ) 12.6
Hz, 1 H, benzyl H), 4.82 (d, J1,2 ) 7.2 Hz, 1 H, 1b-H),4.75-4.70
(m, 3 H, benzyl H), 4.65-4.56 (m, 4 H, 3a-H, benzyl H), 4.54-
4.41 (m, 2 H, benzyl H), 4.30-4.29 (m, 1 H, R-Ser-H), 4.11 (s, 1
H, 4a-H), 3.98 (s, 1 H, 4b-H), 3.83 (s, 2 H, â-Ser-H, â′-Ser-H),
3.77-3.75 (m, 2 H, 2b-H, 5a-H), 3.73 (dd, J6,5 ) J6,6′ ) 9.0, 1 H,
6b-H), 3.68 (s, 2 H, 5b-H, 6a-H), 3.60-3.56 (m, 3 H, 3b-H, 6b′-
H,. 6a′-H), 1.44 (2 × s, 18 H, Boc, t-Bu), 1.04-1.00 (m, 21 H,
TIPS). 13C NMR (151 MHz, CDCl3): δ ) 168.8, 155.6, 139.1-
127.2 (30 C, arom. C), 105.1 (C-1b), 96.7 (C-1a), 84.0 (C-2a), 83.8,
82.0 (C-3b), 80.0, 79.6 (C-2b), 77.3, 75.5 (C-4a), 74.7 (C-3a), 74.7,
74.6, 73.7 (C-4b), 73.6, 73.1, 72.8 (C-5b), 71.8 (C-5a), 69.1 (C-â),
68.2 (C-6a), 68.0, 62.1 (C-6b), 54.1 (C-R), 28.4, 27.9 (2 C),
18.0 (2 C), 11.8 (10 C). MALDI-MS (positive mode, DHB): [M
+ Na]+: m/z calcd.: 1243.6, found: 1242.1. C68H92N2SiO16
(1221.55) calcd.: C: 66.84, H: 7.59, N: 2.29, found: C: 66.04,
H: 6.70, N: 2.56.
O-(-2,3,4,6-Tetra-O-benzyl-â-D-galactopyranosyl)-(1f3)-(2-
acetamido-4-O-benzyl-2-deoxy-6-(triisopropylsilyl)-r-D-galacto-
pyranosyl)-N-(tert-butyloxycarbonyl)-L-serine tert-Butyl Ester
(9a). Following general procedure B from 8a (70 mg, 0.06 mmol)
after flash chromatography with petroleum ether/ethyl acetate (2:
1) compound 9a (51 mg, 0.041 mmol, 69%) was obtained as a
colorless oil. Rf ) 0.25 (petroleum ether/ethyl acetate ) 2:1); [R]D
) +1.4 (c ) 0.1, CHCl3). 1H NMR (600 MHz, CDCl3): δ ) 7.33-
7.18 (m, 25 H, arom. H), 5.51 (d, J ) 8.5 Hz, 1 H, GalNAc-NH),
5.23 (d, J ) 8.4 Hz, 1 H, Ser-NH), 4.97 (d, J ) 11.4 Hz, 2 H,
benzyl H), 4.87 (d, J ) 11.4 Hz, 1 H, benzyl H), 4.86 (br s, 1 H,
1a-H), 4.74 (s, 2 H, 2a-H, benzyl H), 4.69 (d, J ) 12.0 Hz, 1 H,
benzyl H), 4.68 (d, J ) 12.0 Hz, 1 H, benzyl H), 4.54-4.43 (m, 2
H, benzyl H, 1b-H), 4.43-4.40 (m, 3 H, benzyl H), 4.30 (br s, 1
H, R-Ser-H), 4.06 (s, 1 H, 4a-H), 3.94 (s, 1 H, 4b-H), 3.84-3.78
(m, 3 H, 2b-H, 3a-H, â-Ser-H), 3.74-3.69 (m, 4 H, 6a-H, 5a-H,
â′-Ser-H, 6b-H), 3.65-3.53 (m, 4 H, 5b-H, 3b-H, 6b′-H, 6a′-H),
2.04 (s, 3 H, OAc), 1.45, 1.44 (2 × s, 18 H, Boc, t-Bu), 1.04-1.00
(m, 21 H, TIPS). 13C NMR (151 MHz, CDCl3): δ ) 105.8 (C-
1b), 99.0 (C-1a), 82.1 (C-3b), 79.4 (C-2b), 78.5 (C-3a), 75.8 (C-
4a), 73.5 (C-4b), 73.1 (C-5b), 72.0 (C-5a), 68.6 (C-â), 68.3 (C-
6a), 62.9 (C-6b), 54.4 (C-R), 49.1 (C-2a), 29.7-11.8 (14 C).
MALDI-MS (positive mode, DHB): [M + Na]+: m/z calcd.:
1255.6, found: 1256.8. C70H96N2O15Si (1233.60) calcd.: C: 68.16,
H: 7.84, N: 2.27, found: C: 68.16, H: 7.87, N: 2.16.
O-(-2,3,4,6-Tetra-O-benzyl-â-D-galactopyranosyl)-(1f3)-(2-
acetamido-4-O-benzyl-2-deoxy-6-(triisopropylsilyl)-r-D-galacto-
pyranosyl)-N-(tert-butyloxycarbonyl)-L-threonine tert-Butyl Es-
ter (9b). Following the procedure for the synthesis of 8a from 6
(100 mg, 0.11 mmol), 7b (72 mg, 0.25 mmol), and potassium tert-
butoxide in dry toluene after flash chromatography with petroleum
ether/ethyl acetate (2:1) 8b was obtained. From this product
following general procedure B compound 9b (57 mg, 0.046 mmol,
42%) was obtained as a colorless oil. Rf ) 0.23 (petroleum ether/
1
ethyl acetate ) 2:1); [R]D ) +5.5 (c ) 0.45, CHCl3). H NMR
(600 MHz, CDCl3): δ ) 7.39-7.16 (m, 25 H, arom. H), 5.63 (d,
J ) 9.0 Hz, 1 H, GalNAc-NH), 5.07 (d, J ) 9.5 Hz, 1 H, Thr-
NH), 4.92 (d, J ) 11.4 Hz, 2 H, benzyl H), 4.78-4.79(m, 2 H,
1a-H, 2a-H), 4.74-4.66 (m, 3 H, benzyl H), 4.51 (d, J1,2 ) 7.8
Hz, 1 H, 1b-H), 4.52 (d, J ) 10.8 Hz, 2 H, benzyl H), 4.42-4.37
(m, 3 H, benzyl H), 4.10-4.09 (m, 2 H, R-Thr-H, â-Thr-H), 4.04
(s, 1 H, 4a-H), 3.95 (s, 1 H, 4b-H), 3.82-3.70 (m, 4 H, 2b-H,
3a-H, 5a-H, 6a-H), 3.69-3.64 (m, 1 H, 6b-H), 3.55 (d, J ) 9.0
Hz, 3 H, 3b-H, 6b′-H, 6a′-H), 1.72 (s, 3 H, NAc) 1.46, 1.44 (2 ×
s, 18 H, Boc, t-Bu), 1.28 (d, J ) 6.0 Hz, 3 H, Me), 1.02-0.98 (m,
21 H, TIPS). 13C NMR (151 MHz, CDCl3): δ ) 139.1-127.1 (30
C, arom. C), 105.8 (C-1b), 100.4 (C-1a), 82.2 (C-3b), 79.6 (C-2b),
78.6 (C-3a), 76.1 (C-â), 76.0 (C-4a), 74.8, 74.5, 74.1, 73.6 (C-4b),
73.1 (C-5b), 72.3 (C-5a), 71.1, 68.6 (C-6a), 63.2 (C-6b), 58.8 (C-
R), 48.6 (C-2a), 29.7 (3 C), 28.4 (3 C), 28.1 (3 C), 23.4 (2 C), 18.0
(2 C), 11.8 (10 C). MALDI-MS (positive mode, DHB): [M + Na]+:
m/z calcd.: 1269.7, found: 1269.0. C71H98N2O15Si (1247.63)
calcd.: C: 68.35, H: 7.92, N: 2.25, found: C: 68.08, H: 8.00,
N: 2.12.
O-(2,3,4,6-Tetra-O-benzyl-â-D-galactopyranosyl)-(1f3)-4-O-
benzyl-D-galactal (11). To a mixture of TIPS-protected saccharide
5 (320 mg, 0.35 mmol) and THF (20 mL) was added TBAF in
THF (0.4 mL, 1 m solution). After 6 h the solvent was removed.
Purification by flash chromatography (toluene/ethyl acetate ) 5:1)
afforded after lyophilization 11 (225 mg, 0.30 mmol, 86%) as a
colorless powder; mp 43 °C. Rf ) 0.20 (toluene/ethyl acetate )
5:1); [R]D ) -36.8 (c ) 1, CHCl3). 1H NMR (250 MHz, CDCl3):
δ ) 7.38-7.42 (m, 25 H, arom. H), 6.40 (dd, J1,2 ) 6.2 Hz, J1,3
)
1.6 Hz, 1 H, 1a-H), 5.10 (t, J ) 11.8 Hz, J ) 10.7 Hz, 3 H, benzyl
H), 4.84-4.70 (m, 5 H, 2a-H, benzyl H), 4.62-4.51 (m, 3 H, 3a-
H, benzyl H), 4.42 (s, 2 H, 1b-H, benzyl H), 3.96-3.75 (m, 5 H,
4a-H, 5a-H, 4b-H, 2b-H, 6-H), 3.60-3.54 (m, 5 H, 3 × 6-H, 5b-
H, 3b-H), 2.51 (s, 1 H, OH). 13C NMR (63 MHz, CDCl3): δ )
144.5 (C-1a), 138.2-127.7 (30 C, arom. C), 103.1 (C-1b), 100.1
(C-2a), 96.1, 82.1 (C-3b), 79.7 (C-2b), 75.3 (C-5a), 74.6, 73.9 (C-
4b), 73.6, 73.5, 73.2, (C-5b), 72.3 (C-5a), 71.8 (C-3b), 71.2 (C-
4a), 69.0 (C-6a), 62.0 (C-6b). MALDI-MS (positive mode, DHB):
[M + Na]+: m/z calcd.: 781.3, found: 780.9. C47H50O9 (758.89),
calcd.: C: 74.38, H: 6.64, found: C: 74.57, H: 6.73.
O-(2-O-Azido-3,4,6-tri-O-benzyl-2-deoxy-â-D-glucopyranosyl)-
(1f6)-[(O-(2,3,4,6-tetra-O-benzyl-â-D-galactopyranosyl)-(1×a83)]-
4-O-benzyl-D-galactal (12). A mixture of 11 (189 mg, 0.25 mmol)
and 10A[12] (160 mg, 0.27 mmol) was dissolved in MeCN and
cooled to -40 °C, and tin(II) triflate (4 mg, 0.01 eq) in MeCN (1
mL) was added dropwise. After 30 min NaHCO3 and CH2Cl2 were
added and the suspension was washed with water. The solution
was dried with MgSO4 and the solvent removed under reduced
pressure. Purification by flash chromatography (toluene/ethyl acetate
20:1) afforded 12 (182 mg, 0.15 mmol, 60%) as a slightly yellow
oil. Rf ) 0.51 (toluene/ethyl acetate ) 20:1); [R]D ) -10.5 (c )
0.5, CHCl3). 1H NMR (600 MHz, CDCl3): δ ) 7.34-7.14 (m, 40
H, arom. H), 6.40 (d, J1,2 ) 5.6 Hz, 1 H, 1a-H), 5.05 (d, J ) 12.0
Hz, 2 H, benzyl H), 4.98 (d, J ) 10.8 Hz, 1 H, benzyl H), 4.92 (d,
J ) 10. Hz, 1 H, benzyl H), 4.85 (d, J ) 10.7 Hz, 1 H, benzyl H),
4.83 (d, J ) 5.7 Hz, 1 H, 2a-H), 4.78-4.68 (m, 7 H, benzyl H),
4.56 (d, J ) 4.2 Hz, 1 H, 3a-H), 4.53-4.51 (m, 2 H, 1b-H, benzyl
4374 J. Org. Chem., Vol. 72, No. 12, 2007

Mucin Core Structure Synthesis
H), 4.45-4.40 (m, 3 H, benzyl H), 4.23 (br s, 1 H, 1c-H), 4.19 (s,
1 H, 5a-H), 3.84 (dd, J2,1 ) J2,3 ) 8.7 Hz, 1 H, 2b-H), 3.91-3.90
(m, 3 H, 4b-H, 6a-H, 6a′-H), 3.66-3.53 (m, 7 H, 3b-H, 4b-H, 4c-
H, 6b-H, 6b-′H, 6c-H, 6c′-H), 3.54 (d, J ) 5.8 Hz, 2 H, 2c-H,
3c-H), 3.27 (d, J ) 8.4 Hz, 1 H, 5c-H). 13C NMR (151 MHz,
CDCl3): δ ) 144.2 (C-1a), 138.8-127.5 (48 C, arom. C), 102.8
(C-1b), 102.4 (C-1c), 100.1 (C-2a), 83.0 (C-3b), 82.1 (C-3b), 79.6
(C-2b), 77.6 (C-4c), 76.0 (C-5a), 75.4, 75.2, 74.9, 74.9 (C-5c), 74.8,
74.5, 73.8 (C-4b), 73.8, 73.5, 73.4 (C-5b), 73.1, 71.4, (C-3a), 71.4
(C-4a), 69.0 (C-6a), 68.7 (C-6b), 68.4 (C-6c), 66.2 (C-2a). MALDI-
MS (positive mode, DHB): [M + Na]+: m/z calcd.: 1238.5,
found: 1237.5. HRMS (ESI) m/z calcd for [C74H77N3O13 + Na]+
1238.5348, found 1238.5325.
O-(2,3,4,6-Tetra-O-benzyl-r-D-galactopyranosyl)-(1f3)-(2-ac-
etamido-di-4,6-O-benzyl-2-deoxy-r-D-galactopyranosyl)-N-(tert-
butyloxycarbonyl)-L-serine tert-Butyl Ester (22a). Following
general procedure B from 21a (140 mg, 0.12 mmol) after flash
chromatography with petroleum ether/ethyl acetate (5:1), compound
22a (97 mg, 0.083 mmol, 69%) was obtained as a colorless oil. Rf
) 0.22 (toluene/ethyl acetate ) 5:1). [R]D ) +5.1 (c ) 0.3, CHCl3).
1H NMR (600 MHz, CDCl3): δ ) 7.34-7.11 (m, 30 H, arom. H),
5.98 (d, J ) 8.9 Hz, 1 H, GalNAc-NH), 5.47 (d, J ) 8.4 Hz, 1 H,
Ser-NH), 5.05 (d, J ) 11.5 Hz, 1 H, benzyl H), 4.94 (d, J1,2 ) 3.2
Hz, 1 H, 1b-H), 4.85 (d, J ) 11.6 Hz, 2 H, benzyl H), 4.83 (d, J
) 11.8 Hz, 1 H, benzyl H), 4.81 (d, J1,2 ) 4.1 Hz, 1 H, 1a-H),
4.70-4.60 (m, 4 H, 2a-H, benzyl H), 4.53-4.37 (m, 4 H, benzyl
H), 4.29-4.22 (m, 2 H, R-Ser-H, benzyl H), 4.12-4.07 (m, 2 H,
2b-H, 5a-H), 3.95-3.93 (m, 1 H, 3b-H), 3.85-3.77 (m, 6 H, 3a-
H, 4a-H, 4b-H, 5b.H, â-Ser-H, â′-Ser-H), 3.53-3.47 (m, 3 H, 6a-
H, 6b-H, 6b′-H), 3.42-3.40 (m, 1 H, 6a′-H), 1.86 (s, 3 H, Ac),
1.42, 1.40 (2 × s, 18 H, t-Bu, Boc). 13C NMR (63 MHz, CDCl3):
δ ) 170.3, 169.6, 139.2-127.0 (36 C, arom. C), 99.2 (C-1b), 98.9
(C-1a), 82.1, 79.1 (C-3b), 77.6 (C-3a), 76.3 (C-2b), 75.1 (C-4b),
74.8, 74.6, 74.4, 73.8, 73.0, 70.1 (C-5a), 70.0 (C-5b), 69.6 (C-â),
69.5 (C-6a), 68.6 (C-6b), 67.1, 54.5 (C-R), 48.9 (C-2a), 28.3 (4
C), 28.0 (4 C), 23.1 (2 C). MALDI-MS (positive mode, DHB):
[M + Na]+: m/z calcd.: 1189.6, found: 1188.8. C68H82N2O15
(1167.38) calcd.: C: 69.96, H: 7.08, N: 2.40, found: C: 68.94,
H: 7.04, N: 2.27.
O-(2,3,4,6-Tetra-O-benzyl-r-D-galactopyranosyl)-(1f3)-(2-ac-
etamido-4-O-benzyl-2-deoxy-6-O-(triisopropylsilyl)-<bold>R-
D-galactopyranosyl)-N-(tert-butyloxycarbonyl)-L-serine tert-
Butyl Ester (24a). Following general procedure B from 23a (50
mg, 0.043 mmol) after flash chromatography with toluene/ethyl
acetate (5:1), compound 24a (36 mg, 0.031 mmol, 72%) was
obtained as a colorless oil. Rf ) 0.65 (toluene/ethyl acetate ) 5:1);
[R]D ) +11.6 (c ) 1, CHCl3). 1H NMR (250 MHz, CDCl3): δ )
7.21-7.00 (m, 25 H, arom. H), 5.83 (d, J ) 8.8 Hz, 1 H, GalNAc-
NH), 5.20 (d, J ) 8.6 Hz, 1 H, Ser-NH), 4.98 (d, J ) 11.4 Hz, 1
H, benzyl H), 4.81 (d, J1,2 ) 3.6 Hz, 1 H, 1b-H), 4.77-4.68 (m, 2
H, benzyl H), 4.64 (d, J1,2 ) 2.0 Hz, 1 H, 1a-H), 4.52-4.49 (m, 3
H, 2a-H, benzyl H), 4.45-4.22 (m, 5 H, benzyl H), 4.20-4.05 (m,
1 H, R-H), 3.96-3.89 (m, 2 H, 2b-H, 5b-H), 3.83-3.75 (m, 2 H,
3b-H, 4a-H), 3.66-3.51 (m, 7 H, 3a-H, 4b-H, 5a-H, 6a-H, 6a′-H,
â-Ser-H, â′-Ser-H), 3.32-3.24 (m, 2 H, 6b-H, 6b′-H), 1.72 (s, 3
H, Ac), 1.43, 1.40 (2 × s, 18 H, t-Bu, Boc), 1.03-0.90 (m, 21 H,
TIPS). 13C NMR (63 MHz, CDCl3): δ ) 168.6, 163.0, 137.0-
125.3 (30 C, arom. C), 97.6 (C-1b), 97.5 (C-1a), 94.5, 80.1 (C-
3b), 77.8 (C-3a), 76.3 (â-C), 74.5 (C-4a), 73.8 (4b-H), 73.3, 73.0
(C-5a), 72.7, 72.3, 71.5, 70.7 (C-5b), 68.6, 67.8, 67.3, 62.7, 60.5,
53.1, 53.0 (C-R), 47.5 (C-2a), 26.7, 10.3 (16 C). MALDI-MS
(positive mode, DHB): [M + Na]+: m/z calcd.: 1256.6, found:
1257.5. C70H96N2O15Si (1233.60) calcd.: C: 68.15, H: 7.84, N:
2.27, found: C: 67.77, H: 8.35, N: 2.38.
O-(2-O-Azido-3,4,6-tri-O-benzyl-2-deoxy-â-D-glucopyranosyl)-
(1f6)-[(O-(2,3,4,6-tetra-O-benzyl-â-D-galactopyranosyl)-(1×a83)]-
4-O-benzyl-2-nitro-D-galactal (13). Following general procedure
A from 12 (320 mg, 0.35 mmol), after flash chromatography with
toluene/ethyl acetate (10:1) compound 13 (228 mg, 0.24 mmol,
68%) was obtained as a slightly yellow oil. Rf ) 0.38 (toluene/
1
ethyl acetate ) 20:1); [R]D ) +2.5 (c ) 0.25, CHCl3). H NMR
(600 MHz, CDCl3): δ ) 8.13 (s, 1 H, 1a-H), 7.33-7.15 (m, 40 H,
arom. H), 5.24 (br s, 1 H, 3a-H), 4.97 (d, J ) 11.5 Hz, 2 H, benzyl
H), 4.93 (d, J1,2 ) 7.6 Hz, 1 H, 1b-H), 4.83 (d, J ) 10.8 Hz, 1 H,
benzyl H), 4.73-4.70 (m, 3 H, benzyl H), 4.69-4.67 (m, 1 H,
4a-H), 4.62 (d, J ) 9.1 Hz, 1 H, benzyl H), 4.58-4.40 (m, 9 H,
benzyl H), 4.11 (d, J ) 8.3 Hz, 1 H, 6a-H), 4.06 (d, J1,2 ) 6.9 Hz,
1 H, 1c-H), 3.90-3.88 (m, 3 H, 4b-H, 5a-H, 6a′-H), 3.78 (dd, J1,2
) J2,3 ) 10.2 Hz, 1 H, 2b-H), 3.67-3.69 (m, 1 H, 5b-H), 3.29-
3.56 (m, 5 H, 3b-H, 4c-H, 6b-H, 6c-H, 6c′-H), 3.52 (d, J ) 9.7
Hz, 1 H, 6b′-H), 3.39 (dd, J2,1 ) J2,3 ) 6.9 Hz, 1 H, 2c-H), 3.26
(dd, J3,4 ) J3,2 ) 6.9 Hz, 1 H, 3c-H) 3.13 (d, J ) 7.8 Hz, 1 H,
5c-H). 13C NMR (151 MHz, CDCl3): δ ) 155.1 (C-1a), 138.8-
127.5 (48 C, arom. C), 103.5 (C-1b), 102.6 (C-1c), 82.8 (C-3c),
82.2 (C-3b), 79.8 (C-2b), 78.7 (C-4a), 77.3 (C-4c), 75.4, 75.2, 75.1,
74.9 (C-5c), 74.8, 74.7, 74.2 (C-4b), 73.4, 73.3 (C-5b), 72.8, 72.5,
72.1, 70.7 (C-5a), 68.8 (C-6c), 68.3 (C-6a), 68.0 (C-6b), 66.1 (C-
2a). MALDI-MS (positive mode, DHB): [M + Na]+: m/z calcd.:
1283.5, found: 1282.5. HRMS (ESI) m/z calcd for [C74H76N4O15
+ Na]+ 1283.5199, found 1283.5178.
O-(2-O-Azido-3,4,6-tri-O-benzyl-2-deoxy-â-D-glucopyranosyl)-
(1f6)-[(O-(2,3,4,6-tetra-O-benzyl-â-D-galactopyranosyl)-(1f3)]-
4-O-benzyl-2-deoxy-2-nitro-r-D-galactopyranosyl)-N-(tert-bu-
tyloxycarbonyl)-L-serine tert-Butyl Ester (14a). To a mixture of
13 (40 mg, 0.032 mmol), 7a (12 mg, 0.048 mmol), and dry toluene
(1 mL) was added potassium tert-butoxide (0.4 mg, 0.003 mmol)
under stirring. After 2 h the reaction mixture was neutralized with
acetic acid and the solvent concentrated under reduced pressure.
Purification by flash chromatography (toluene/ethyl acetate ) 10:
1) afforded 14 (31 mg, 0.0020 mmol, 63%) as a colorless oil. Rf )
0.55 (toluene/ethyl acetate ) 20:1); [R]D ) +0.6 (c ) 0.08, CHCl3).
1H NMR (600 MHz, CDCl3): δ ) 7.34-7.14 (m, 40 H, arom. H),
6.40 (d, J1,2 ) 4.6 Hz, 1 H, 1a-H), 5.23 (d, J ) 8.8 Hz, 1 H, Ser-
NH), 5.00 (d, J ) 11.4 Hz, 1 H, benzyl H), 4.91 (dd, J2,1 ) 4.6
Hz, J2,3 ) 9.6 Hz, 1 H, 2a-H), 4.87-4.85 (m, 2 H, benzyl H), 4.78
(m, 3 H, 1b-H, benzyl H), 4.72 (d, J ) 13.4 Hz, 1 H, benzyl H),
4.64-4.53 (m, 8 H, 3a-H, benzyl H), 4.46-4.44 (m, 3 H, benzyl
H), 4.29-4.26 (m, 2 H, 1c -H, R-Ser-H), 4.00 (br s, 1 H, 4a-H),
3.96 (br s, 2 H, 4b-H, 5a-H), 3.90 (br s, 1 H, â-Ser-H), 3.83 (d, J
) 9.1 Hz, 1 H, â-H), 3.78-3.77 (m, 1 H; 2b-H), 3.74-3.55 (m, 9
H, 3b-H, 4c-H, 5c-H, 6b-H, 6b′-H, 6c-H, 6c′-H, 6a-H, 6a′-H), 3.38-
3.35 (m, 2 H, 5b-H, 3c-H), 3.31 (dd, J2,1 ) J2,3 ) 8.2 Hz, 1 H,
2c-H), 1.47, 1.44 (2′ s, 18 H, Boc, t-Bu). 13C NMR (151 MHz,
CDCl3 (selected data)): δ ) 105.0 (C-1b), 102.2 (C-1c), 96.5 (C-
1a), 83.8 (C-2a), 83.0 (C-3c), 82.1 (C-3b), 79.5 (C-2b), 77.6 (C-
4c), 75.6 (C-4a), 74.9 (C-5b), 73.8 (C-4b), 72.9 (C-5c), 70.1 (C-
5a), 69.1 (C-â), 68.5 (C-6b), 68.2 (C-6c), 66.5 (C-2c), 54.3 (C-R).
MALDI-MS (positive mode, DHB): [M + Na]+: m/z calcd.:
1544.7, found: 1544.2. HRMS (ESI) m/z calcd for [C86H99N5O20
+ Na]+ 1544.6775, found 1544.6765.
O-(2,3,4,6-Tetra-O-benzyl-r-D-galactopyranosyl)-(1f3)-(2-ac-
etamido-4-O-benzyl-2-deoxy-6-O-(triisopropylsilyl)-<bold>R-
D-galactopyranosyl)-N-(tert-butyloxycarbonyl)-L-threonine tert-
Butyl Ester (24b). Following general procedure B from 23b (50
mg, 0.04 mmol) after flash chromatography with toluene/ethyl
acetate (5:1), compound 24b (35 mg, 0.03 mmol, 70%) was
obtained as a colorless oil. Rf ) 0.69 (toluene/ethyl acetate ) 5:1);
1
[R]D ) +2.9 (c ) 0.19, CHCl3). H NMR (600 MHz, CDCl3): δ
) 7.3-7.19 (m, 25 H, arom. H), 6.00 (d, J ) 8.9 Hz, 1 H, GalNAc-
NH), 5.24 (d, J ) 8.2 Hz, 1 H, Thr-NH), 5.10 (d, J ) 12.0 Hz, 1
H, benzyl H), 4.97 (d, J1,2 ) 3.6 Hz, 1 H, 1b-H), 4.88 (d, J ) 12.0
Hz, 1 H, benzyl H), 4.80 (d, J ) 11.0 Hz, 1 H, benzyl H), 4.76 (s,
2 H, 1a-H, 2a-H), 4.69-4.62 (m, 2 H, benzyl H), 4.50 (d, J )
12.0 Hz, 1 H, benzyl H), 4.40 (t, J ) 11.4 Hz, 1 H, benzyl H),
4.38-4.23 (m, 3 H, benzyl H), 4.13-4.10 (m, 3 H, 3b-H, R-Thr-
H, â-Thr-H), 4.06 (dd, J2,1 ) 3.6 Hz, J2,3 ) 9.6 Hz, 1-H, 2b-H),
3.98 (s, 1 H, 3b-H), 3.96 (s, 1 H, 4a-Η,), 3.81-3.77 (m, 3 H, 3a-
H, 4b-H, 5a-H), 3.72-3.69 (m, 2 H, 6a-H, 6a′-H), 3.48-3.47 (m,
J. Org. Chem, Vol. 72, No. 12, 2007 4375

Geiger et al.
1 H, 6b-H), 3.43-3.40 (m, 1 H, 6b′-H), 1.93 (s, 3 H, Ac), 1.42,
1.38 (2 × s, 18 H, t-Bu, Boc), 1.26 (d, J ) 8.4 Hz, 3 H, Me),
1.03-1.00 (m, 21 H, TIPS). 13C NMR (151 MHz, CDCl3, selected
data): δ ) 100.3 (C-1a), 98.6 (C-1b), 79.4 (C-3b), 77.3 (C-3a),
76.1 (C-â), 76.3 (2b-H), 75.5 (4b-H), 74.3 (C-4a), 72.4 (C-5a),-
70.1 (C-5b), 69.3 (C-6b), 62.3 (C-6a), 58.6 (C-R), 48.7 (C-2a).
MALDI-MS (positive mode, DHB): [M + Na]+: m/z calcd.:
1269.6, found: 1268.5. HRMS (ESI) m/z calcd for [C71H98N2O15-
Si + Na]+ 1269.6629, found 1269.6593.
73.4, 72.6 (C-5a), 71.4, 70.6 (C-5b), 68.8 (C-6b), 68.6 (C-6a), 68.5
(â-C), 56.9 (C-2a), 54.4 (R-C), 48.7 (C-2b). MALDI-MS (positive
mode, DHB): 1141.1 [M + Na]+. Anal. Calcd for C63H79N3O15
(1117.55): C, 67.66; H, 7.12; N, 3.76, found: C, 67.72; H, 7.17;
N, 3.81.
O-(2-Actamido-2-deoxy-3,4,6-tri-O-benzyl-â-D-glucopyrano-
syl)-(1f6)-(3,4-di-O-benzyl-2-acetamido-2-deoxy-D-galactopy-
ranosyl)-N-(tert-butyloxycarbonyl)-L-threonine tert-Butyl Ester
(35b). 35b was prepared in 79% yield from 34b following the same
procedure as for the preparation of 35a. [R]D ) +10.5 (c ) 0.25,
O-(2-Azido-2-deoxy-3,4,6-tri-O-benzyl-â-D-glucopyranosyl)-
(1f3)-(4-O-benzyl-2-deoxy-2-nitro-6-O-(triisopropylsilyl)-r-D-
galactopyranosyl)-N-(tert-butyloxycarbonyl)-L-serine tert-Butyl
Ester (28a). To a solution of 27 (150 mg, 0.17 mmol) and 7a (66
mg, 0.25 mmol) in dry toluene (5 mL) was added potassium tert-
butoxide (2 mg, 0.018 mmol) under stirring; after 2 h the mixture
was neutralized with acetic acid and the solvent removed in vacuo.
Purification by flash chromatography (petroleum ether/ethyl acetate
) 10:1) furnished 28a (141 mg, 0.12 mmol, 72%) as a colorless
oil. Rf ) 0.65 (petroleum ether/ethyl acetate ) 3:1); [R]D (c ) 0.2,
1
CHCl3). H NMR (600 MHz, CDCl3): δ ) 7.37-7.18 (m, 25 H,
aromatic), 5.65 (d, J ) 9.9 Hz, 1 H, -NHAc), 5.46 (d, J ) 8.0 Hz,
1 H, -NHAc), 5.05 (d, J ) 9.0 Hz, 1 H, -NHBoc), 4.93 (d, J )
11.4 Hz, 1 H, -OCH2Ph), 4.81-4.75 (m, 3 H, 1a-H, -OCH2Ph),
4.72 (dd, J ) 10.2, 3.6 Hz, 1 H, 2a-H), 4.69-4.63 (m, 3 H, 1b-H,
-OCH2Ph), 4.60-4.54 (m, 3 H, -OCH2Ph), 4.47-4.45 (m, 2 H,
-OCH2Ph), 4.15 (d, J ) 9.0 Hz, 1 H, R-Thre-H), 4.05 (br d, J )
5.4 Hz, 1 H, â-Thre-H), 3.92-3.84 (m, 4 H, 4a-H, 6a-H, 3b-H,
5b-H), 3.74-3.67 (m, 3 H, 6b-H, 6b′-H, 4b-H), 3.65-3.62 (m, 1
H, 6a′-H), 3.56-3.51 (m, 3 H, 2b-H, 3a-H, 5a-H), 1.85 (s, 3 H,
-NHAc), 1.79 (s, 3 H, -NHAc), 1.47, 1.41 (s, 18 H, tert-butyl),
1.22 (d, J ) 5.4 Hz, 3 H, γ-Thre-H). 13C NMR (151 MHz,
CDCl3): δ ) 170.5, 170.0, 155.7, 138.5-127.5 (m, 30 C, aromatic),
100.4 (C-1b), 99.9 (C-1a), 82.7, 80.9 (C-4a), 80.1, 78.2 (C-4b),
77.5 (C-3a), 76.1, 75.9 (â-C), 74.7 (C-5a), 74.5, 74.2, 73.4, 72.8
(C-3b), 71.6, 70.3 (C-5b), 68.7 (C-6b), 68.4 (C-6a), 58.5 (R-C),
56.2 (C-2b), 48.8 (C-2a), 29.7, 28.3, 28.1, 23.5, 18.6. MALDI-MS
(positive mode, DHB): 1155.5 [M + Na]+. Anal. Calcd for
C64H81N3O15 (1131.57): C, 67.88; H, 7.21; N, 3.71, found: C,
68.01, H, 7.28; N, 3.82.
1
CHCl3) ) +3.1. H NMR (600 MHz, CDCl3): δ ) 7.33-7.17
(m, 20 H, arom. H), 5.30 (d, J1,2 ) 3.8 Hz, 1 H, 1a-H), 5.19 (d, J
) 8.6, 1 H, NH), 4.99 (dd, J2,1 ) 3.8 Hz, J2,3 ) 10.5 Hz, 1 H,
2a-H), 4.95 (d, J ) 11.3 Hz, 1 H, benzyl H), 4.88 (d, J ) 11.0 Hz,
1 H, benzyl H), 4.87 (d, J ) 10.7 Hz, 1 H, benzyl H), 4.80-4.77
(m, 3 H, 1b-H, benzyl H), 4.63-4.59 (m, 3 H, 3a-H, benzyl H),
4.55 (d, J ) 13.9 Hz, 1 H, benzyl H), 4.31 (br s, 1 H, R-Ser-H),
4.21 (s, 1 H, 4a-H), 3.84 (d, J ) 6.8 Hz, 4 H, 5a-H, 6b-H, â-Ser-
H, â′-Ser-H), 3.76-3.67 (m, 4 H, 4b-H, 6a-H, 6a′-H, 6b′-H), 3.56-
3.55 (m, 1 H, 5b-H), 3.45 (dd, J3,2 ) J3,4 ) 9.3 Hz, 1 H, 3b-H),
3.32 (dd, J2,1 ) J2,3 ) 9.3 Hz, 1 H, 2b-H), 1.45 (2 × s, 18 H, Boc,
t-Bu), 1.03-1.00 (m, 21 H, TIPS). 13C NMR (151 MHz, CDCl3):
δ ) 137.9-127.3 (24 C, arom. C), 10.7 (C-1b), 99.6 (C-1b), 88.5,
83.6 (C-2a), 82.6 (C-3b), 77.3 (C-4b), 76.0 (C-4a), 75.6 (C-3a),
75.3, 74.8 (C-5b), 73.5, 71.6 (C-5a), 69.0 (C-â), 68.9 (C-6b), 66.4
(C-2b), 61.9 (C-6a), 54.1 (C-R), 28.3, 27.8, 18.0, 11.8 (17 C).
MALDI-MS (positive mode, DHB): [M + Na]+: m/z calcd.:
1178.6, found: 1179.7. C61H85N5O15Si (1156.44). The mass peak
could not be obtained by ESI HRMS.
O-(2-Acetamido-3,4,6-tri-O-acetyl-2-deoxy-r-D-galactopyra-
nosyl)-(1f6)-(2-acetamido-3,4-di-O-acetyl-2-deoxy-r-D-galacto-
pyranosyl)-N-(fluorenylmethyloxycarbonyl)-L-serine tert-Butyl
Ester (42a). 41a (0.15 g, 0.122 mmol) was dissolved in ethanol (5
mL) and transferred to a hydrogenation vessel. Platinized Raney
nickel T4 catalyst was freshly prepared and the material obtained
from 2 g of Raney nickel/aluminum alloy was suspended in ethanol
(15 mL). From a homogeneous suspension of this catalyst 8 mL
were added to the reaction vessel and the mixture shaken in a Parr
apparatus at a hydrogen pressure of 75 psi for 12 h. The catalyst
was filtered off and the solvent evaporated. The residue was
dissolved in pyridine/acetic anhydride (2:1, 6 mL) and stirred for
12 h. All volatiles were removed and the crude product purified
by flash chromatography (toluene/acetone 2:1) to give the inter-
mediate bis-acetamido product (0.108 g, 70%). This material was
dissolved in THF (2 mL) and treated with tetrabutylammonium
fluoride solution (0.1 mL of a 1 M solution in THF) and stirred at
rt for 4 h. The solution was acidified by addition of acetic acid and
concentrated. The crude material was purified by flash chroma-
tography (toluene/acetone 3:2) to give the 6-O-unprotected product,
which was directly used for subsequent debenzylation. The material
was dissolved in methanol/acetic acid (2.5 mL, 4:1) and stirred
over Pd/C (0.02 g, 10% palladium) under hydrogen atmosphere
for 14 h. The catalyst was filtered off, and all volatiles were
removed. The residue was treated with pyridine/acetic anhydride
(2 mL, 2:1) for 12 h. Removal of all volatiles and column
chromatographic purification of the residue (chloroform/methanol
22:1) gave the Boc-protected tert-butyl ester of the target molecule
as a colorless foam (0.0602 g, 56% over five steps). Rf ) 0.54
(chloroform/methanol 13:1); [R]D25 ) +101.3 (c ) 1, CHCl3). 1H
O-(2-Actamido-2-deoxy-3,4,6-tri-O-benzyl-â-D-glucopyrano-
syl)-(1f6)-(3,4-di-O-benzyl-2-acetamido-2-deoxy-D-galactopy-
ranosyl)-N-(tert-butyloxycarbonyl)-L-serine tert-Butyl Ester (35a).
Following general procedure B from 34a (150 mg, 0.116 mmol)
after flash chromatography with petroleum ether/ethyl acetate (5:
1), compound 35a (160 mg, 82%) was obtained as a colorless solid.
34a (150 mg, 0.116 mmol) was dissolved in ethanol (6 mL) and
transferred to a hydrogen vessel. Platinized Raney nickel T4 catalyst
(freshly prepared, and the material obtained from 2 g of Raney
nickel/aluminum alloy was suspended in ethanol, 10 mL) was added
to the reaction vessel and the mixture shaken in a Parr apparatus at
a hydrogen pressure of 75 psi for 24 h. The catalyst was carefully
filtered off and the solvent evaporated. The residue was dissolved
in pyridine/acetic anhydride (2:1, 4 mL) and stirred overnight and
concentrated, and the crude product was purified by flash chro-
matography (chloroform/methanol 20:1) to give compound 6 as a
colorless solid (160 mg, 82%), mp. 204 °C. Rf 0.56 (toluene/acetone
15:1). [R]D ) +24.5 (c ) 0.5, CHCl3). 1H NMR (600 MHz,
CDCl3): δ ) 7.37-7.18 (m, 25 H, aromatic), 5.84 (d, J ) 7.2 Hz,
1 H, -NHAc), 5.47 (d, J ) 7.2 Hz, 1 H, NHBoc), 5.36 (d, J ) 7.8
Hz, 1 H, -NHAc), 4.90 (d, J ) 11.4 Hz, 1 H, -OCH2Ph), 4.81-
4.76 (m, 4 H, 1a-H, -OCH2Ph), 4.74 (d, J ) 7.2 Hz, 1 H, 1b-H),
4.68-4.60 (m, 4 H, 2b-H, -OCH2Ph), 4.57-4.42 (m, 3 H, -OCH2-
Ph), 4.24 (br s, 1 H, R-Ser-H); 4.06 (br s, 1 H, 3a-H), 3.90 (br s,
1 H, 5a-H), 3.89-3.78 (m, 4 H, 6b-H, 6b′-H, 5b-H, â-Ser-H), 3.73-
3.64 (m, 4 H, 6a-H, 6a′-H, â′-Ser-H, 4b-H), 3.57-3.53 (m, 2 H,
3b-H, 4a-H), 3.42 (br t, J ) 9.0 Hz, 1 H, 2a-H), 1.88 (s, 3 H,
-NHAc), 1.84 (s, 3 H, -NHAc), 1.45, 1.42 (2 s, 18 H, tert-butyl).
13C NMR (151 MHz, CDCl3): δ ) 170.3, 169.8, 155.3, 138.5-
127.5 (m, 30C, aromatic), 100.1 (C-1b), 98.8 (C-1a), 82.4, 80.4
(C-3a), 80.1, 78.5 (C-4b), 77.1 (C-3b), 75.0, 74.6 (C-4a), 74.5, 74.2,
3
NMR (600 MHz, CDCl3): δ ) 6.34 (d, JNH,2 ) 9.3 Hz, 1 H,
GalNHAcb-NH), 5.79 (d, 3JNH,R ) 6.3 Hz, 1 H, Ser-NH), 5.66 (d,
3
3JNH,2 ) 9.6 Hz, 1 H, GalNHAca-NH), 5.40 (d, J4,3 ) 3.3 Hz, 1
3
3
H, 4b-H), 5.37 (d, J4,3 ) 3.3 Hz, 1 H, 4a-H), 5.21 (dd, J3,2
)
11.3, 3J3,4 ) 2.3 Hz, 1 H, 3b-H), 5.13 (dd, 3J3,2 ) 11.3, 3J3,4 ) 3.2
3
3
Hz, 1 H, 3a-H), 4.86 (d, J1,2 ) 3.6 Hz, 1 H, 1a-H), 4.81 (d, J1,2
) 3.4 Hz, 1 H, 1b-H), 4.66 (ddd, 3J2,NH ) 10.5, 3J2,1 ) 3.4, 3J2,3
)
)
3
3
3
10.5 Hz, 1 H, 2b-H), 4.62 (ddd, J2,NH ) 9.9, J2,1 ) 3.3, J2,3
9.9 Hz, 1 H, 2a-H), 4.42 (br s, 1 H, R-H), 4.23-4.18 (m, 2 H,
4376 J. Org. Chem., Vol. 72, No. 12, 2007

Mucin Core Structure Synthesis
5b-H, 6b-H), 4.08-3.97 (m, 3 H, 5a-H, 6b′-H, â-H), 3.78-3.74
(m, 2 H, 6a-H, â′-H), 3.31 (br s, 1 H, 6a′-H), 2.16-1.98 (7 s, 21
H, 5 OAc, 2 NHAc), 1.15, 1.46 (2 s, 18 H, 2 C4H9). 13C NMR
(150.8 MHz, CDCl3): δ ) 170.9-170.0 (9 C), 155.2, 97.8 (1a-
C), 97.4 (1b-C), 82.8, 80.1, 68.4 (3b-C), 68.3 (3a-C), 68.0 (â-C),
67.7 (4a-C), 67.6 (4b-C), 67.1 (5b-C), 65.3 (6a-C), 62.1 (6b-C),
54.0 (R-C), 47.8 (2a-C), 47.3 (2b-C), 28.3 (3 C), 28.0 (3 C), 23.3
(2 C), 23.0, 20.7 (4 C). MS (FAB): calcd.: 877 + 1 (H) ) 878,
877 + 23 (Na) ) 900, 877 + 173 (Na, NaI) ) 1050; found 878
[M + H]+, 900 [M + Na]+, 1050 [M + NaI + Na]+. This
intermediate was immediately transformed into the target mole-
cule 42a:
(s, 1 H, 4a-H), 5.27 (d, J ) 8.6 Hz, 1 H, Ser-NH), 5.07 (d, J )
10.8 Hz, 1 H, benzyl H), 4.95 (d, J1,2 ) 2.2 Hz, 1 H, 1a-H), 4.85-
4.83 (m, 1 H, 2b-H), 4.80 (s, 1 H, 1b-H), 4.70-4.65 (m, 2 H, benzyl
H), 4.48 (d, J ) 10.9 Hz, 1 H, benzyl H), 4.25 (br s, 2 H, 2a-H,
R-Ser-H), 4.03 (s, 2 H, 5a-H, 6b′-Η), 3.89-3.85 (m,2 H, 5a-H,
6b′-H), 3.76-3.70 (m, 4 H, 3a-H, 5b-H, 6b′-H, â-Ser-H, â′-Ser-
H), 3.65-3.60 (m, 2 H, 6a′′-H, 6b′′-H), 3.45 (d, J ) 10.3 Hz, 1 H,
3b-H), 2.14, 2.06 (3 × s, 9 H, OAc), 1.47, 1.40 (2 × s, 18 H, t-Bu,
Boc), 1.03-0.95 (m, 30 H, TIPS, t-Bu). 13C NMR (151 MHz,
CDCl3) δ ) 171.2, 170.6 (C-Ac), 138.8-127.2 (24 C, arom. C),
102.4 (C-1b), 98.5 (C-1a), 82.5, 78.2 (C-3b), 77.1 (C-3b), 73.1 (C-
4b), 72.2 (C-5b), 69.9 (C-5a), 69.6 (C-â), 61.4 (C-6b), 61.0 (C-
6a), 54.4 (C-R), 49.6 (C-2a), 49.2 (C-2b), 29.7, 28.3, 28.0, 27.0,
23.1, 23.1, 21.2, 19.1, 17.9, 11.8 (20 C). MALDI-MS (positive
mode, DHB): [M + Na]+: m/z calcd.: 1308.5, found: 1306.9.
C69H101N3O16Si2 (1284.72), calcd.: C: 64.51, H: 7.92, N: 3.27,
found: C: 64.17, H: 8.08, N: 3.02.
The intermediate (0.0120 g, 0.0139 mmol) was dissolved in a
mixture of trifluoroacetic acid and dichloromethane (2 mL, 1:1)
and the solution stirred at rt for 12 h. Then all solvents were
evaporated. The residue was redissolved together with Fmoc-ONSu
(0.0094 g, 0.0278 mmol) and sodium bicarbonate (0.0233 g, 0.2780
mmol) in acetonitrile/water (3 mL, 1:1) and the mixture stirred for
12 h. The solution was diluted with dichloromethane and acidified
with 2 N HCl solution (25 mL) and the aqueous layer extracted
with dichloromethane (3 × 25 mL). The combined organic extracts
were dried over sodium sulfate and the volatiles evaporated. Column
chromatographic purification (10 g silica gel, chloroform/methanol/
acetic acid 8:2:1) of the residue afforded 42a as a colorless foam.
(0.0131 g, quantitative). Rf ) 0.23 (chloroform/methanol 4:1););
[R]D ) +88.8 (c ) 1, CHCl3). 1H NMR (600 MHz, CDCl3): δ )
7.89 (d, J ) 7.5 Hz, 2 H, arom. H), 7.71 (t, J ) 7.7 Hz, 2 H, arom.
O-(3,6-Di-O-benzyl-4-O-tert-butyldimethylsilyl-2-deoxy-2-ni-
tro-â-D-glucopyranosyl)-(1f6)-(3,4-di-O-tert-butyldimethylsilyl-
2-deoxy-2-nitro-r-D-galactopyranosyl)-N-(tert-butyloxycarbonyl)-
l-serine tert-Butyl Ester (55a). 54 (0.044 g, 0.049 mmol) and 7a
(0.026 g, 0.098 mmol) were carefully dried under high vacuum
and dissolved in dry toluene. The reaction was activated by addition
of potassium tert-butoxide solution (49 µL of a 1 m solution in
THF) at rt under constant stirring. After 45 min the reaction was
quenched with acetic acid and concentrated. Flash chromatography
of this material (toluene/ethyl acetate 20:1) furnished 55a as a
colorless oil (0.028 g, 49%). Rf ) 045 (toluene/ethyl acetate 10:
1); [R]D25 ) +50.9 (c ) 2, CHCl3). 1H NMR (600 MHz, CDCl3):
δ ) 7.33-7.22 (m, 10 H, arom. H), 5.39 (d, 3JNH,R ) 8.8 Hz, 1 H,
3
H), 7.40-7.30 (m, 4 H, arom. H), 5.45 (d, J4,3 ) 3.2 Hz, 1 H,
3
4a-H), 5.33 (d, J4,3 ) 2.6 Hz, 1 H, 4b-H), 5.21-5.17 (m, 2 H,
3
3
3a-H, 3b-H), 4.88 (d, J1,2 ) 3.3 Hz, 1 H, 1a-H), 4.77 (d, J1,2
)
3
3
3.5 Hz, 1 H, 1b-H); 4.55-4.42 (m, 4 H, â-H, â′-H, 2a-H, 2b-H),
4.35 (t, 3J5,6 ) 6.6 Hz, 3J5,6′ ) 5.6 Hz, 1 H, 5a-H), 4.28-4.23 (m,
3 H, 5b-H, R-H, Fmoc-CH), 4.02 (dd, 3J6,5 ) 6.0, 2J6,6′ ) 11.2 Hz,
NH), 5.18 (d, J1,2 ) 4.1 Hz, 1 H, 1a-H), 5.03 (d, J1,2 ) 8.2 Hz,
1 H, 1b-H), 4.84 (dd, 3J2,1 ) 4.1, 3J2,3 ) 10.3 Hz, 1 H, 2a-H), 4.67
2
2
(d, J ) 10.7 Hz, 1 H, benzyl H), 4.60 (d, J ) 12.2 Hz, 1 H,
benzyl H), 4.52 (dd, 3J2,1 ) 8.2, 3J2,3 ) 10.0 Hz, 1 H, 2b-H), 4.50
(d, 2J ) 12.2 Hz, 1 H, benzyl H), 4.45 (d, J ) 10.7 Hz, 1 H, benzyl
3
2
1 H, 6b-H), 3.95 (s, 2 H, Fmoc-CH2), 3.83 (dd, J6′,5 ) 7.0, J6,6′
3
2
) 11.1 Hz, 1 H, 6b′-H), 3.77 (t, J6,5 ) 9.6, J6,6′ ) 9.6 Hz, 1 H,
3
2
3
3
6a-H), 3.34 (dd, J6′,5 ) 5.0, J6′,6 ) 9.9 Hz, 1 H, 6a′-H), 2.14-
1.86 (7 s, 21 H, 5 OAc, 2 NHAc). 13C NMR (150.8 MHz, CDCl3):
δ ) 174.0-172.0 (8 C), 145.3-121-0 (12 C), 100.0 (1a-C), 99.1
(1b-C), 71.5 (Fmoc-CH2),70.1 (3a-C), 69.8 (3b-C), 69.3 (4a-C),
68.8 (2 C, 5a-C, 4b-C), 68.2 (â-C), 68.0 (5b-C), 67.4 (6a-C), 63.0
(6b-C), 57.7 (Fmoc-CH), 49.0 (2a-C), 48.7 (2b-C), 48.4 (R-C), 22.8,
22.7, 20.7 (2 C), 20.6 (2 C), 20.5. MS (FAB): calcd. 943 + 23
(Na) ) 966, 943 + 46 (2 Na)- 1 (H) ) 988; found 966 (M +
Na)+, 988 (M + 2 Na-H)+. HRMS (ESI) m/z calcd for [C44H53N3O20
+ Na]+ 966.3114, found 966.3116.
O-(2-Acetamido-3,4-di-O-benzyl-2-deoxy-6-O-(tert-butyldiphe-
nylsilyl)- r-D-galactopyranosyl)-(1f3)-(4-O-acetyl-2-acetamido-
2-deoxy-6-O-(triisopropylsilyl)-r-D-galactopyranosyl)-N-(tert-
butyloxycarbonyl)-L-serine tert-Butyl Ester (47a). To a solution
of 46a (40 mg, 0.03 mmol) in ethanol (5 mL) was added freshly
prepared platinized Raney Nickel T4 catalyst, then stirred and left
to react for 12 h under hydrogen atmosphere at 50 bar. The catalyst
was filtered off and purification by flash chromatography (toluene/
ethyl acetate ) 2:1) furnished 47a (17 mg, 0.008 mmol, 26%) and
as a second product 6-O-TBDPS-protected 40a (9 mg, 0.01 mmol,
33%) as a colorless lyophilisate. 47a: Rf ) 0.35 (toluene/ethyl
acetate 2:1); [R]D (c ) 0.12, CHCl3) ) +11.8. 1H NMR (600 MHz,
CDCl3): δ ) 7.63 (d, J ) 6.7 Hz, 2 H, arom. H) 7.62 (d, J ) 6.7
Hz, 2 H, arom. H), 7.41-7.24 (m, 16 H, arom. H), 6.83 (d, J )
8.6 Hz, 1 H, b-GalNAc-NH), 5.77 (s, 1 H, a-GalNAc-NH), 5.42
H), 4.39 (dd, J3,2 ) 10.3, J3,4 ) 2.4 Hz, 1 H, 3a-H), 4.24 (m, 1
H, R-H), 4.10 (t, 3J3,2 ) 9.1, 3J3,4 ) 9.1 Hz, 1 H, 3b-H), 3.97 (dd,
3JR,â ) 2.9, 3Jâ,â ) 11.2 Hz, 1 H, â-H), 3.85-3.40 (m, 2 H, 5a-H,
6a-H), 3.81 (s, 1 H, 4a-H), 3.78-3.75 (m, 2 H, 4b-H, â-CH), 3.70-
3.69 (m, 2 H, 6a′-H, 6b-H), 3.64 (dd, 3J6,5 ) 4.7, 2J6,6′ ) 10.9 Hz,
1 H, 6b′-H), 3.60-3.59 (m, 1 H, 5b-H), 1.49, 1.45 (2 s, 18 H, 2
C4H9), 0.89-0.83 (3 s, 27 H, 3 C4H9), 0.25-0.00 (6 s, 18 H, 6
SiCH3). 13C NMR (151 MHz, CDCl3): δ ) 169.2, 155.4, 137.9,
136.9, 128.3-127.5 (10 C), 99.3 (1b-C), 97.5 (1a-C), 89.9 (2b-C),
85.6 (2a-C), 82.6 (3b-C), 82.3 (2-C), 79.8 (2-C), 76.6 (5b-C), 74.9,
73.3, 70.7 (4a-C), 70.3 (â-C), 69.9 (5a-C), 68.9 (3a-C), 68.1 (6b-
C), 66.0 (6a-C), 54.4 (R-C), 28.4, 27.9, 26.1, 25.8, 18.5, 18.1, 17.9,
-2.2, -3.5, -4.9, -5.6 (2 C). MS (FAB): calcd.: 1165 + 23
(Na) ) 1188; found 1188 [M + Na]+. HRMS (ESI) m/z calcd for
[C56H95N3O17Si + Na]+ 1188.5861, found 1188.5870.
Acknowledgment. This work was supported by the Deutsche
Forschungsgemeinschaft and the Fonds der Chemischen
Industrie.
Supporting Information Available: NMR spectra, experimen-
tal procedures, and physical data. This material is available free of
charge via the Internet at http://pubs.acs.org.
JO061670B
J. Org. Chem, Vol. 72, No. 12, 2007 4377