Convergent Synthesis of Sialyl LewisX- O-Core-1 Threonine

Article abstract of DOI:10.1021/acs.joc.7b03117

Selectins are a class of cell adhesion molecules that play a critical role during the initial steps of inflammation. The N-terminal domain of P-selectin glycoprotein ligand-1 (PSGL-1) binds to all selectins, but with the highest affinity to P-selectin. Re

Full text of DOI:10.1021/acs.joc.7b03117

Subscriber access provided by UNIV OF SCIENCES PHILADELPHIA
Convergent synthesis of sialyl LewisX-O-Core-1 threonine
Mohammed Y. R. Sardar, Appi Reddy Mandhapati, Simon Park,
Walter J. Wever, Richard D. Cummings, and Elliot L. Chaikof
J. Org. Chem., Just Accepted Manuscript • DOI: 10.1021/acs.joc.7b03117 • Publication Date (Web): 11 Apr 2018
Downloaded from http://pubs.acs.org on April 11, 2018
Just Accepted
“Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted
online prior to technical editing, formatting for publication and author proofing. The American Chemical
Society provides “Just Accepted” as a service to the research community to expedite the dissemination
of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in
full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully
peer reviewed, but should not be considered the official version of record. They are citable by the
Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore,
the “Just Accepted” Web site may not include all articles that will be published in the journal. After
a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web
site and published as an ASAP article. Note that technical editing may introduce minor changes
to the manuscript text and/or graphics which could affect content, and all legal disclaimers and
ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or
consequences arising from the use of information contained in these “Just Accepted” manuscripts.
is published by the American Chemical Society. 1155 Sixteenth Street N.W.,
Washington, DC 20036
Published by American Chemical Society. Copyright © American Chemical Society.
However, no copyright claim is made to original U.S. Government works, or works
produced by employees of any Commonwealth realm Crown government in the course
of their duties.

Page 1 of 30
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
Convergent synthesis of sialyl LewisXꢀOꢀCoreꢀ1 threonine
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Mohammed Y. R. Sardar†,‡, Appi Reddy Mandhapati†,‡, Simon Park†,‡, Walter J. Wever†,‡,
Richard D. Cummings†, and Elliot L. Chaikof†,‡,§*
†Department of Surgery, Center for Drug Discovery and Translational Research, Beth Israel
Deaconess Medical Center, Harvard Medical School, 110 Francis Street, Suite 9F, Boston,
Massachusetts 02215, USA.
‡Wyss Institute of Biologically Inspired Engineering, Harvard University, 110 Francis Street,
Suite 9F, Boston, Massachusetts 02115, USA.
§HarvardꢀMIT Division of Health Sciences and Technology, Cambridge, MA 02139
Submitted as a Revised Research Article to
The Journal of Organic Chemistry
joꢀ2017ꢀ03117y
*Corresponding author
Elliot L. Chaikof, M.D., Ph.D.
Department of Surgery
Beth Israel Deaconess Medical Center
110 Francis St, Suite 9F
Boston, MA 02115
Eꢀmail: echaikof@bidmc.harvard.edu
Phone: 617ꢀ632ꢀ9581
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 2 of 30
2
1
2
3
4
Abstract
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Selectins are a class of cell adhesion molecules that play a critical role during the initial steps
of inflammation. The Nꢀterminal domain of Pꢀselectin glycoprotein ligandꢀ1 (PSGLꢀ1) binds to all
selectins, but with highest affinity to Pꢀselectin. Recent evidence suggests that the blockade of Pꢀ
selectin/PSGLꢀ1 interactions provides a viable therapeutic option for the treatment of many
inflammatory diseases. Herein, we report the total synthesis of threonine bearing sialyl LewisX (sLeX)
linked to a Coreꢀ1ꢀOꢀhexasaccharide 1, as a key glycan of the Nꢀterminal domain of PSGLꢀ1. A
convergent synthesis using αꢀselective sialylation and a regioselective [4+2] glycosylation are the key
features of this synthesis.
ACS Paragon Plus Environment

Page 3 of 30
The Journal of Organic Chemistry
3
1
2
3
4
Introduction
5
6
7
8
During the course of inflammation, homing of leukocytes to vascular endothelial cells and
subsequent extravasation represents critical initial features of the evolving immune response.1ꢀ3 The
homing of leukocytes is governed by the interactions between cell adhesion molecules and their
ligands.1,4,5 Pꢀ, Lꢀ, and Eꢀselectin represent a class of Ca2+ dependent cell adhesion receptors that are
expressed on activated endothelial cells, platelets, and leukocytes. Pꢀselectin glycoprotein ligandꢀ1
(PSGLꢀ1), which is expressed on leukocytes, binds to all three selectins with highest affinity towards
Pꢀselectin, followed by Lꢀ and Eꢀselectin.6,7 The interaction of PSGLꢀ1 on leukocytes and Pꢀselectin
on endothelial cells initiates a cascade of events that results in the rolling, arrest, and subsequently,
the transendothelial migration of leukocytes. Similarly, the interaction of PSGLꢀ1 on leukocytes and Pꢀ
selectins on activated platelets initiates the formation of plateletꢀleukocyte aggregates that facilitates
the infiltration of leukocytes at sites of injury.1 Persistent upregulation of selectins leads to excessive
trafficking of leukocytes, which may result in tissue damage that underlies a wide range of chronic
disorders of innate immunity including atherosclerosis, inflammatory bowel disease, and metabolic
syndrome.8 For these reasons, the blockade of selectin/ligand interactions holds therapeutic potential
for these and other conditions including thrombosis and cancer.9
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
The role of Pꢀselectin/PSGLꢀ1 interaction in various inflammatory diseases has motivated the
investigation of a number of approaches to block Pꢀselectin/PSGLꢀ1 interactions, albeit with modest
success. For instance, while antibody based therapies appear promising, manufacturing cost and the
development of antibodies against these biologics limits their effectiveness.10 There have also been
numerous attempts to design small molecule inhibitors by modifying sialyl LewisX (sLeX), a critical
binding motif, which has yielded a series of low affinity inhibitors including GMIꢀ1070,11,12 PSIꢀ697,13,14
and TBC1269.15 The interaction between PSGLꢀ1 and Pꢀselectin is governed not only by sLeX, but by
the presence of a neighboring cluster of tyrosine sulfates and the amino terminal peptide
backbone.16,17 Consequently, a number of efforts have been directed towards the synthesis of the Nꢀ
terminal region of PSGLꢀ1 as a tool compound for drug discovery.
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 4 of 30
4
1
2
3
4
5
6
7
8
9
The generation of glycosulfopeptides that mimic PSGLꢀ1 remains limited, in part, by the lack
of a scalable chemical synthesis for sLeXꢀCoreꢀ1ꢀThr. More specifically, the absence of a facile and
scalable synthesis of the sLeX tetrasaccharide and the Coreꢀ1 disaccharide are the key bottlenecks
for the generation of sLeXꢀCoreꢀ1ꢀThr. While we have developed a scalable process to generate
Coreꢀ1 diol in multiꢀgram scale,18 the design of a scheme for the efficient, large scale synthesis of
sLeX that can be coupled to Coreꢀ1 has remained an elusive synthetic target despite attempts by
several research groups over past two decades.19ꢀ30 For example, initial reports from Nicolaou19 and
Danishefsky29 described the synthesis of sLeX in 7 to 9 steps with overall yields ranging from 5% to
20%. More recently, Boons and coworkers reported the synthesis of sLeX derivative in 7 steps with
35% overall yield, but on a 35 mg scale.22 Key challenges of sLeX synthesis include the requirement
of an αꢀ(2→3) sialylation and αꢀ(1→3) fucosylation, along with the acid lability of fucose, and the poor
reactivity of glucosamine derivatives.31ꢀ35 Moreover, glycosylation of sLeX donors offers a unique set
of challenges. In nature, all sLeX derivatives are βꢀlinked through an Nꢀacetyl glucosamine C1.
However, glycosylation using unmodified Nꢀacetyl glucosamine donors leads to an inert oxazoline36,37
and the glycosylation involving a sLeX donor and a Coreꢀ1 diol acceptor is associated with poor
regioselectivity. Notably, only two synthetic schemes for C2ꢀOꢀsLeX hexasaccharide have been
reported and both are limited by either an absence of control over regioꢀ or diastereoselectivity.22,38 In
particular, the synthesis of a similar structure reported by Kunz and coworkers suffers a major
setback at the key [4+2] glycosylation step resulting in an undesired 1→4 linked hexasaccharide as
the major product.38 While improved regioselectivity was observed on a modified mimic of sialyl
LewisX39, significant formation of undesired 1→4 linkage continued to be a major flaw, emphasizing
the need for an improved method.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Recently, we have synthesized an analogue of the Nꢀterminal domain of PSGLꢀ1, GSnPꢀ6,
which binds to human Pꢀselectin with a Kd of 22 nM.18 In this approach, we utilized
glycosyltransferases to add the terminal galactose, sialic acid, and fucose. In order to produce GSnPꢀ
6, as well as other glycosulfopeptide targets of biological interest in large scale, we herein report the
ACS Paragon Plus Environment

Page 5 of 30
The Journal of Organic Chemistry
5
1
2
3
4
5
6
7
8
9
total chemical synthesis of a threonine bearing C2ꢀOꢀsLeX hexasaccharide 1 (Fig. 1). Our synthetic
scheme takes advantage of αꢀselective sialylation using 4ꢀO, 5ꢀNꢀoxazolidone protection. An iterative
αꢀselective sialylation forms the desired disaccharide donor with minimum modification followed by
convergent synthesis of sLeX based on [2+2] glycosylation and subsequent coupling to the Coreꢀ1
diol.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Fig. 1. Sialyl LewisXꢀOꢀCoreꢀ1 threonine 1.
Results and Discussion
We envisioned that [2+2] glycosylation to generate sLeX with subsequent [4+2]
glycosylation between the sLeX donor and a Coreꢀ1 diol acceptor would be the most efficient
route to afford C2ꢀOꢀsLeX (Fig. 2). This approach would allow us to address the challenges
posed by αꢀ(1→3) fucosylation and αꢀ(2→3) sialylation during the generation of relatively less
complex disaccharides 9 and 16.
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 6 of 30
6
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Fig. 2. Retrosynthesis of hexasaccharide 1.
We began our investigation with a focus on αꢀ(2→3) sialylation, which has often
impeded the large scale generation of sLeX (Scheme 1). NꢀAcetyl oxazolidone protected
sialoside donors, reported by Crich and coworkers, exhibit excellent αꢀselectivity.40,41
Subsequently, Wong and coworkers extended the utility of 5ꢀN,4ꢀOꢀoxazolidone protection
towards sialoside phosphates for the αꢀselective iterative glycosylation. As a consequence, αꢀ
(2→3) sialylation of the galactosyl acceptor 4 was pursued using sialoside phosphate donor 742
to generate αꢀ(2→3) SiaꢀGal disaccharide 8 in one step. The synthesis of galactosyl acceptor 4
was produced from commercially available D
ꢀgalactose, as reported elsewhere.43 The
commercially available sialoside donor 5 was converted to the oxazolidone protected
thioglycoside 6 in two steps. Subsequently, the thioglycoside was converted to its phosphate
donor 7 by glycosylation using NIS/TfOH and dibutylphosphate. Iterative glycosylation was
ACS Paragon Plus Environment

Page 7 of 30
The Journal of Organic Chemistry
7
1
2
3
4
5
6
7
8
9
carried out in a 3:1 mixture of dichloromethane and acetonitrile at ꢀ70°C using TMSꢀOTf. The
formation of an αꢀ(2→3) linkage was confirmed by the 3JC1,Hꢀ3ax coupling constant (5.5 Hz) and
the chemical shift of Hꢀ3eq (2.99 ppm), which were in agreement with prior reports.44,45
Acetylation of the disaccharide using pyridine and Ac2O resulted in the final disaccharide donor
9.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Scheme 1. Synthesis of sialylgalactose disaccharide 9.
With successful preparation of the sialic acidꢀgalactose disaccharide donor 9, we
focused on the preparation of the glucosamineꢀfucose disaccharide acceptor 16 (Scheme 2).
Anomeric OTBDPS46 and NHTroc47 protecting groups were selected due to their compatibility
with the subsequent synthesis of more complex analogues. Our initial approach was to
fucosylate the diol acceptor 14 regioselectively, as reported by Magnusson and coworkers27 for
a similar acceptor. We sought to bypass regioselective ring opening of 4,6ꢀOꢀbenzylidene
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 8 of 30
8
1
2
3
4
5
6
7
8
acetal, which requires the use of a strong acid in the presence of an acid labile fucose. We
initially synthesized Trocꢀprotected glucosamine 12, followed by deacetylation under Zémplen
conditions to afford triol, which was converted in situ to 4,6ꢀOꢀ benzylidene acetal 13 by
9
treatment with PhCH(OMe)2 and CSA to afford the 3ꢀOH acceptor.46 Regioselective cleavage of
the benzylidene acetal was carried out using triethylsilane and TFA to afford the diol 14.
Glycosylation between 14 and fucose donor 15 was carried out using TMSꢀOTf at ꢀ15°C in
CH2Cl2 to obtain the disaccharide 16. However, the purification of the desired
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Scheme 2. Synthesis of disaccharide 16.
(1→3) linked disaccharide from the undesired (1→4) linked disaccharide was challenging,
particularly at large scale. As an alternative approach, fucosylation of acceptor 13 was
performed followed by regioselective cleavage of the 4,6ꢀOꢀbenzylidene acetal. Glycosylation
between 13 and 15 was carried out using TMSꢀOTf at ꢀ20°C in CH2Cl2. Purification of the
resulting disaccharide from excess donor was carried out after regioselective cleavage of
48
benzylidene acetal using TfOH and TES at ꢀ78°C in CH2Cl2 . While other choices were
available, the TES/TfOH system was selected to cleave the 4,6ꢀOꢀbenzylidene acetal, since the
ACS Paragon Plus Environment

Page 9 of 30
The Journal of Organic Chemistry
9
1
2
3
4
5
6
7
8
9
reaction could be carried out at ꢀ78°C and cleavage of fucose was not observed. The structure
of disaccharide 16 was confirmed by 1D and 2D NMR spectroscopy (Supporting Information).38
Sialyl LewisX tetrasaccharide 17 was generated by convergent [2+2] glycosylation
between sialylgalactosyl thioglycoside donor 9 and fucosylglucosamine acceptor 16 (Scheme
3). The glycosylation was carried out using a NIS/TfOH activation system at ꢀ40°C in CH2Cl2,
which proceeded smoothly to form fully protected sLeX 17 in 86% yield. NMR spectroscopy
revealed that the isolated compound consisted solely of the βꢀanomeric configuration, which
was confirmed by the presence of a large coupling constant (JH1ꢀH2 = 8.4 Hz). To the best of our
knowledge, this is the shortest synthesis of sLeX reported to date. The synthesis of over one
gram of fully protected sLeX was accomplished in 5 steps from the monosaccharide building
blocks, with 51% overall yield.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Scheme 3. Synthesis of sialyl LewisX.
Subsequently, sLeX 17 was treated with TBAF and catalytic AcOH to remove the
OTBDPS protecting group at the anomeric OH, which was then converted to the imidate donor
18 in two steps using CCl3CN and DBU in 64% yield. The key [4+2] glycosylation between the
sLeX imidate donor and the Coreꢀ1 diol was carried out using TMSꢀOTf catalyst in CH2Cl2
(Scheme 4). However, the resulting product consisted of a 3:1 mixture of the desired 1→6 and
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 10 of 30
10
1
2
3
4
5
6
7
8
9
the undesired 1→4 linkages. Despite the evaluation of alternate reaction conditions, significant
improvement was not observed and attempts to conduct the reaction below ꢀ20°C were
unsuccessful. This limitation of regioselectivity involving a donor with a participating group and
the 4,6ꢀdiol system has been previously reported by other groups.38,49,50 While it is unexpected
for the axial 4ꢀOH to react, as proposed by Baumann, it is possible that the 6ꢀOH reacts with the
intermediate oxazolinium ion via the exocyclic carbon and upon intramolecular rearrangement
forms the 1→4 linked product (Fig. 3).51 To circumvent this limitation, we postulated that a
thioglycoside donor would enable glycosylation at a much lower temperature, which would
potentially improve regioselectivity. Hence, the imidate donor was converted to its thioglycoside
19 by treatment with TMSꢀOTf and thiophenol in one step with 82% yield.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Fig. 3. Speculated mechanism for the formation of the 1→4 linkage.
Table 1 summarizes the results of [4+2] glycosylation to a Coreꢀ1 diol acceptor using
two sLeX donors and different promoters under various conditions (Table 1). As noted, the sLeX
imidate donor 18 produced a 3:1 mixture of products upon activation using catalytic TMSꢀOTf in
CH2Cl2. Use of a less polar solvent, such as a 3:1 mixture of CH2Cl2 and toluene, did not
improve regioselectivity and further attempts to lower the reaction temperature led to the
absence of product, while a higher temperature resulted in the loss of fucose. Studies with the
sLeX thioglycoside donor 19 were initially conducted at ꢀ40°C using NIS and TfOH. Higher yield
and an improved regioselectivity (10:1) favoring the βꢀ(1→6) linked hexasaccharide was
observed (Table 1, entry 3). Given the pronounced effect of reaction temperature on selectivity
and overall yield, we explored conditions under which reaction temperature was further reduced.
ACS Paragon Plus Environment

Page 11 of 30
The Journal of Organic Chemistry
11
1
2
3
4
5
6
7
8
9
Glycosylation was carried out at ꢀ60°C using BSPꢀTf2O along with a bulky base, TTBP (Table 1,
entry 4). Although only a single isomer was isolated, yield was significantly reduced, which may
reflect assisted transfer of a proton by TTBP from glucosamine NHTroc to form an inert
oxazoline.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Scheme 4. [4+2] Glycosylation between sLeX donors and Coreꢀ1 diol acceptor. i a Reaction conditions
and yields are listed in Table 1.
Table 1. Conditions for the [4+2] glycosylation between sLeX donors and Coreꢀ1 diol acceptor.
Yield
Entry
Donor
Promoter
TMSꢀOTf
TMSꢀOTf
NIS/TfOH
Solvent
CH2Cl2
Temperature
ꢀ15°C
(Selectivity)a
67%
(3:1)
1
2
3
4
5
sLeXꢀimidate
sLeXꢀimidate
CH2Cl2/Toluene
CH2Cl2
ꢀ15°C
54%
(3:1)
sLeXꢀthioglycoside
ꢀ40°C
82%
10:1
sLeXꢀthioglycoside BSP/Tf2O/TTBP
sLeXꢀthioglycoside
BSP/Tf2O
CH2Cl2
ꢀ60°C
46%
1→6 only
86%
1→6 only
CH2Cl2
ꢀ60°C
aRatio of 1→6 to 1→4 linked hexasaccharides
The highest yield, with exclusive formation of the βꢀ(1→6) linked hexasaccharide, was
observed when [4+2] glycosylation was carried out with a BSPꢀTf2O promoter in the absence of
a base (Table 1, entry 5). Structural assignments of the final hexasaccharide were performed by
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 12 of 30
12
1
2
3
4
5
6
7
8
a combination of 1H, 13C, COSY, HSQC, TOCSY, and HMBC (Supporting Information). A
downfield 13C shift of CI6, HMBC correlation between CIIIꢀ1 and HIꢀ6, and JH1ꢀH2 confirmed
formation of the βꢀ(1→6) linkage.
9
In conclusion, several key challenges have been addressed in the synthesis of a
threonine bearing C2ꢀOꢀsLeX hexasaccharide. The reported scheme, which describes
stereoselective sialylation, provides an efficient synthesis for sLeX in high yield. Specifically,
convergent synthesis of sLeX in five steps was performed in gram scale with elimination of the
formation of any undesired regioꢀ and stereoisomers. Furthermore, conditions were optimized
for the regioselective [4+2] glycosylation between sLeX and coreꢀ1 diol. A sLeX thioglycoside
enabled low temperature glycosylation using a BSPꢀTf2O activation system. As a complex
naturally occurring glycan, the synthesis of the C2ꢀOꢀsLeX hexasaccharide provides an
opportunity for the creation of tool compounds to probe molecular mechanisms relevant to the
development of therapeutic compounds of clinical interest.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Experimental Section
General
All reactions were performed under inert atmosphere of nitrogen or argon, unless
otherwise noted. DꢀGalactose, Dꢀglucosamine hydrochloride, and Lꢀfucose were purchased
(Carbosynth LLC, CA). All other reagents were purchased from commercial sources and used
directly. All solvents were dried and distilled following standard protocols. All glycosylation
reactions were performed in oven dried round bottom flask. Proton nuclear magnetic resonance
(1H NMR) and 13C NMR spectra were recorded with a Varian 400ꢀMHz and Bruker 600ꢀMHz
spectrometer. High Resolution Mass spectra (HRMS) were acquired using UltraFlex II
MALDI/TOF Mass Spectrometer (Bruker Corporation, MA) and Orbitrap Lumos MS (Thermo
Fisher Scientific, CA). Thin layer chromatography (TLC) was performed on silica gel matrix with
ACS Paragon Plus Environment

Page 13 of 30
The Journal of Organic Chemistry
13
1
2
3
4
5
6
a 254 nm fluorescent indicator and flash column chromatography purification was performed on
Silica Gel 60 (SigmaꢀAldrich Corporation, WI).
7
8
Phenyl 6ꢀOꢀbenzylꢀ1ꢀthioꢀ βꢀDꢀgalactopyranoside (4)
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
TFA (5.0 mL, 64.9 mmol) and triethylsilane (11.0 mL, 68.8 mmol) were added to a
solution of galactose diol 343 (4.60 gm, 12.8 mmol) in CH2Cl2 (50 mL) at 0°C and stirred at room
temperature for 3 h. The reaction was monitored by TLC and upon completion of the reaction,
concentrated in vacuo. The crude was dissolved in EtOAc (100 mL), washed with satd aq
NaHCO3 (2 x 50 mL), brine (100 mL), dried over Na2SO4, filtered, and concentrated in vacuo.
The crude residue was purified by column chromatography over silica gel using 60% EtOAc in
hexanes to obtain a white solid (4.07 gm, 88 %). [α]D24 ꢀ32.6 (c = 1.0, CHCl3), Rf 0.35
(hexane/EtOAc, 1:4); 1H NMR (400 MHz; CD3OD) δ: 7.58 – 7.50 (m, 2H, Aromatic.), 7.35 – 7.14
(m, 8H, Aromatic.), 4.62 (d, J = 9.7 Hz, 1H, Hꢀ1), 4.52 (s, 2H, CH2Ph), 3.87 (d, J = 3.3 Hz, 1H,
Hꢀ4), 3.76 – 3.68 (m, 3H, Hꢀ5, 6ꢀCH2), 3.62 (t, J = 9.4 Hz, 1H, Hꢀ2), 3.51 (dd, J = 9.2, 3.4 Hz,
1H, Hꢀ3); 13C NMR (100.5 Hz, CD3OD) δ: 138.3, 134.7, 130.6, 130.6, 128.5, 127.9, 127.4,
127.2, 126.5, 88.7, 88.6, 77.8, 77.7, 74.8, 73.0, 72.9, 72.9, 69.8, 69.4, 69.3; ESIꢀHRMS m/z
calcd for C19H22O5SNa [M + Na]+ 385.1086, found 385.1074.
Methyl (phenyl 5ꢀacetamidoꢀ7,8,9ꢀtriꢀOꢀacetylꢀ5ꢀN,4ꢀOꢀcarbonylꢀ3,5ꢀdideoxyꢀ2ꢀthioꢀ
Dꢀ
glyceroꢀβꢀ ꢀgalactoꢀnonꢀ2ꢀulopyranoside)onate (6)
D
Sialoside thioglycoside 5 (6.25 g, 10.7 mmol) in methanol (100 mL) was treated with
methanesulfonic acid (2.1 mL, 32.1 mmol) and refluxed under N2 for 24 h. The reaction mixture
was quenched with trimethylamine (10 mL) and then concentrated under reduced pressure. The
contracted residue and NaHCO3 (4.5 g, 53.5 mmol) were dissolved in CH3CN (30 mL) and H2O
(60 mL), and cooled to 0°C. 4ꢀNitrophenyl chloroformate (5.4 g, 26.7 mmol) in CH3CN (30 mL)
was slowly added to the vigorously stirred solution. The reaction mixture was stirred for 3 h at
0°C, after which the aqueous layer was extracted with ethyl acetate (3x100 mL) and the
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 14 of 30
14
1
2
3
4
5
6
combined organic layer was washed with brine, dried over Na2SO4, and concentrated. The
residue was purified by column chromatography over silica gel eluting with 10% methanol in
7
8
9
ethyl acetate to afford 6 as a white foam (3.16 gm, 74%). The NMR spectra for 6 matched those
previously reported.40,41 1H NMR (400 MHz, CDCl3) δ: 7.48 (d, J = 7.6 Hz, 2H), 7.38ꢀ7.28 (m,
3H), 6.99 (br s, 1H), 4.65 (t, J = 11.6, 1H), 4.56 (d, J = 11.2 Hz, 1H), 4.02 (br s, 1H), 3.92 (br s,
1H), 3.83ꢀ3.66 (m, 3H), 3.59 (s, 3H), 3.55 (t, J = 10.4, 1H), 2.85 (d, J = 10.0 Hz, 1H), 2.55 – 2.38
(m, 2H), 2.33 (t, J = 12.4 Hz, 1H); 13C NMR (100.5 Hz, CDCl3) δ: 168.8, 160.7, 136.3, 130.1,
129.0, 128.2, 89.0, 77.6, 75.2, 75.1, 71.2, 70.2, 63.4, 58.4, 58.3, 53.0, 52.9, 36.9.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Dibutyl[(methyl 5ꢀacetamidoꢀ7,8,9ꢀtriꢀOꢀacetylꢀ5ꢀN,4ꢀOꢀcarbonylꢀ3,5ꢀdideoxyꢀ
α/βꢀ ꢀgalactoꢀnonꢀ2ꢀulopyranosyl)onate] phosphate (7)
Acetic anhydride (30 mL) and DMAP (0.61 gm, 5.0 mmol) were added to a solution of
Dꢀglyceroꢀ
D
sialoside thioglycoside 6 (2.1 gm, 5.30 mmol) in pyridine (30 mL) and stirred for 48 h. The
solvent was then removed and the residue was dissolved in CH2Cl2 (100 mL) and successively
washed with an aqueous solution of NaHCO3 (3x30 mL), brine, dried over Na2SO4, and
concentrated in vacuo. The residue was filtered through a silica gel column using 70%
ethylacetate in hexanes. The filtrate was concentrated to obtain a white foam. The white foamy
residue, dibutylphosphate (2.58 mL, 12.8 mmol) and activated 4 Å molecular sieves (3.1 gm)
were stirred in CH2Cl2 (50 mL) for 2 h before cooling to ꢀ10°C followed by the addition of NIS
(1.9 gm, 8.3 mmol) and TfOH (44 ꢁL, 0.5 mmol). The reaction mixture was stirred for 4 h and
was then quenched by the addition of trimethylamine (0.3 mL), diluted in CH2Cl2 (50 mL), and
filtered through a pad of Celite. The filtrate was successively washed with saturated aqueous
solution of Na2S2O3 (30 mL), NaHCO3 (30 mL), brine, dried over Na2SO4, and concentrated in
vacuo. The residue was purified by column chromatography over silica gel using 50% EtOAc in
hexanes to obtain 7 as a yellowish foam (2.74 mg, 82 %). The NMR spectra for 7 matched
those previously reported.42 1H NMR (400 MHz, CDCl3) δ: (αꢀisomer) 5.64 (dd J = 8.0, 0.8 Hz,
ACS Paragon Plus Environment

Page 15 of 30
The Journal of Organic Chemistry
15
1
2
3
4
5
6
7
8
9
1H), 5.30ꢀ5.25 (m, 1H), 4.73 (dt, J = 1.2, 9.6 Hz, 1H), 4.37 (dd, J = 12.3, 2.8 Hz, 1H), 4.17ꢀ4.03
(m, 6H), 3.85ꢀ3.82 (m, 4H), 2.96 (dd, J = 4.4, 12.4 Hz, 1H), 2.64 (t, J = 12.7 Hz, 1H), 2.45 (s,
3H), 2.47 (s, 3H), 2.11 (s, 3H), 2.07 (s, 3H), 2.00 (s, 3H), 1.67ꢀ1.63 (m, 4H), 1.42ꢀ1.36 (m, 4H),
0.90 (t, J = 7.9 Hz, 6H); (βꢀisomer): 5.63 (dd J = 1.2, 4.0 Hz, 1H), 5.23ꢀ5.18 (m, 1H), 4.69 (dd, J
= 2.0, 9.6 Hz, 1H), 4.54 (m, 1H), 4.15 (dd, J = 7.2, 12.4 Hz, 1H), 4.10ꢀ4.05 (m, 4H), 3.82 (s, 3H),
3.73 (dd, J = 9.8, 11.2 Hz, 1H), 2.85 (dd, J = 3.6, 12.8 Hz, 1H), 2.47(s, 3H), 2.27 (dt, J = 2.4,
12.8 Hz, 1H) 2.08 (s, 3H), 2.06 (s, 3H), 1.99 (s, 3H), 1.66ꢀ1.61 (m, 4H), 1.39ꢀ1.35 (m, 4H), 0.90
(t, J = 7.4 Hz, 6H) 13C NMR (100.5 Hz, CDCl3) δ: (αꢀisomer):171.7, 170.5, 170.4, 170.0, 169.9,
167.2, 167.2, 153.4, 98.8, 98.1, 74.1, 71.5, 69.8, 68.1, 68.0, 62.5, 58.3, 53.4, 35.9, 32.1, 32.0,
24.5, 20.9, 20.8, 20.6, 18.6, 18.5, 13.4; (βꢀisomer): 172.1, 170.5, 170.4, 169.8, 169.6, 165.5,
153.4, 98.8, 98.7, 76.6, 73.9, 72.5, 71.7, 68.5, 68.4, 68.3, 62.8, 58.8, 53.4, 36.0, 32.1, 32.0,
24.6, 20.9, 20.7, 18.5, 13.5.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Methyl 5ꢀacetamidoꢀ7,8,9ꢀtriꢀOꢀacetylꢀ5ꢀN,4ꢀOꢀcarbonylꢀ3,5ꢀdideoxyꢀDꢀglyceroꢀαꢀDꢀ
galactoꢀnonꢀ2ꢀulopyranosylonateꢀ(2→3)ꢀphenyl 6ꢀOꢀbenzylꢀthioꢀβꢀDꢀ
galactothiopyranoside (8)
Sialoside phosphate donor 7 (700 mg, 1.1 mmol) and galactosyl thioglycoside acceptor 4
(506 mg, 1.4 mmol), along with activated 4 Å molecular sieves (400 mg) were added to a
mixture of CH2Cl2 and CH3CN (3:1, 20 mL) and stirred for 2 h at room temperature. The mixture
was then cooled to ꢀ78°C and stirred for an additional 30 min, before addition of TMSꢀOTf (200
ꢁL, 1.1 mmol). After stirring for 30 min at ꢀ70°C, the reaction mixture was quenched with Et3N,
(0.5 mL), diluted with CH2Cl2, and filtered through a pad of Celite. The filtrate was successively
washed with water, brine, dried over Na2SO4, concentrated in vacuo, and the resulting residue
was purified by column chromatography over silica gel using 60% EtOAc in hexanes to obtain 8
as a white foam (758 mg, 84%). [α]D24 ꢀ42.7 (c = 1.0, CHCl3), Rf 0.40 (hexane/EtOAc, 1:3); 1H
NMR (400 MHz, CDCl3) δ: 7.60ꢀ7.57 (m, 2H), 7.34ꢀ7.23 (m, 8H), 5.68 (dd, J = 1.6, 8.2 Hz, 1H,
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 16 of 30
16
1
2
3
4
5
6
7
8
9
Hꢀ7’), 5.49 (dt, J = 3.2, 6.8 Hz, 1H, Hꢀ8’), 4.71 (d, J = 9.6 Hz, 1H, Hꢀ1), 4.57ꢀ4.54 (m, 3H, Hꢀ6a’,
PhCH2), 4.37 (dd, J = 3.2, 12.0 Hz, 1H, Hꢀ9’), 4.11 (dd, J = 3.0, 9.6 Hz, 1H, Hꢀ3), 4.02 (dd, J =
6.4, 12.4 Hz, 1H, Hꢀ9b’), 3.96 (dd, J = 2.8, 10.8 Hz, 1H, Hꢀ4’), 3.81ꢀꢀ3.68 (m, 9H, CH3, Hꢀ2, Hꢀ4,
Hꢀ5, Hꢀ6a, Hꢀ6b, Hꢀ5’), 2.99 (dd, J = 3.6, 12.4 Hz, 1H, Hꢀ3e), 2.81 (d, J = 1.6 Hz, 1H), 2.63 (d, J
= 3.2 Hz, 1H), 2.49 (s, 3H, OAc), 2.22 (t, J = 12.8 Hz, 1H, Hꢀ3a), 2.12 (s, 3H, OAc), 2.11 (s, 3H,
OAc), 1.97 (s, 3H, OAc); 13C NMR (100.5 Hz, CDCl3) δ: 171.9, 170.7, 170.1, 169.8, 168.2 (3JCꢀ1,
Hꢀ3ax = 5.5 Hz, Cꢀ1’), 153.4, 137.9, 133.0, 132.2, 128.7, 128.4, 127.3, 127.6, 137.5, 98.1 (Cꢀ2’),
87.7 (Cꢀ1), 78.7 (Cꢀ3), 76.7 (Cꢀ6’), 74.7, 73.6, 71.5 (Cꢀ8’), 69.4, 68.8 (Cꢀ9’), 68.5, 68.4, 67.5,
63.1 (Cꢀ9’), 58.9, 53.2 (Cꢀ1’OOCH3), 36.3 (Cꢀ3’), 29.6, 24.6, 21.2, 20.9, 20.7; HRꢀMALDIꢀ
ToF/MS m/z calcd for C38H45NNaO17S [M + Na]+ 842.2306, found 842.2341.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Methyl 5ꢀacetamidoꢀ7,8,9ꢀtriꢀOꢀacetylꢀ5ꢀN,4ꢀOꢀcarbonylꢀ3,5ꢀdideoxyꢀDꢀglyceroꢀαꢀDꢀgalactoꢀ
nonꢀ2ꢀulopyranosylonateꢀ(2→3)ꢀphenyl 2,6ꢀdioꢀOꢀacetylꢀ6ꢀOꢀbenzylꢀ1ꢀthioꢀβꢀDꢀ
galactothiopyranoside (9)
The disaccharide 8 (720 mg, 0.88 mmol) was dissolved in pyridine (8 mL) followed by the
addition of Ac2O (5 mL) and DMAP (40 mg, 0.33 mmol), which was stirred at room temperature
for 12 h. The reaction mixture was concentrated and diluted in CH2CL2 (50 mL), washed with
saturated aqueous NaHCO3, brine, dried over Na2SO4, filtered, and concentrated in vacuo. The
crude residue was purified by column chromatography over silica gel using 50% EtOAc in
hexanes to obtain a white foam (738 mg, 93%). [α]D24 ꢀ38.2 (c = 1.0, CHCl3), Rf 0.46
(hexane/EtOAc, 1:1); 1H NMR (400 MHz, CDCl3) δ: 7.55ꢀ7.51 (m, 2H), 7.32ꢀ7.22 (m, 8H), 5.64ꢀ
5.62 (m, 1H, Hꢀ7’, Hꢀ8’), 5.14 (t, J = 9.6 Hz, 1H, Hꢀ2), 5.00 (d, J = 3.2 Hz, 1H, Hꢀ4), 4.91 (d, J =
10.0 Hz, 1H, Hꢀ1), 4.62 (dd, J = 3.2, 10.0 Hz, 1H, Hꢀ3), 4.51ꢀ4.45 (m, 3H, Hꢀ9a’,PhCH2), 4.26
(dd, J = 1.4, 9.4 Hz, 1H, Hꢀ6’), 3.99ꢀ3.91 (m, 2H, Hꢀ9b’, Hꢀ5), 3.86 (dd, J = 3.0, 13.2 Hz, 1H, Hꢀ
4’), 3.82 (s, 3H, CO2Me), 3.64 (dd, J = 9.4, 10.6 Hz, 1H, Hꢀ5’), 3.54 (dd, J = 6.2, 10.2 Hz, 1H, Hꢀ
6a), 3.45 (dd, J = 6.2, 10.2 Hz, 1H, Hꢀ6b), 2.92 (dd, J = 3.4, 11.8 Hz, 1H, Hꢀ3e’), 2,45 (s, 3H,
ACS Paragon Plus Environment

Page 17 of 30
The Journal of Organic Chemistry
17
1
2
3
4
5
6
OAc), 2.24 (s, 3H, OAc), 2.13 (s, 3H, OAc), 2.07 (s, 3H, OAc), 2.05 (s, 3H, OAc), 1.85 (t, J =
12.4 Hz, 1H, Hꢀ3a’), 1.85 (s, 3H, OAc); 13C NMR (100.5 Hz, CDCl3) δ: 171.6, 170.4, 170.2,
7
8
9
169.8, 169.4, 167.4, 153.4, 137.9, 133.4, 131.8, 128.7, 128.3, 127.7, 127.4, 97.5 (Cꢀ2’), 85.3 (Cꢀ
1), 75.8, 75.6, 74.6, 73.4, 73.3, 72.8, 72.1, 68.5, 68.4, 68.2, 68.0, 67.9, 63.3, 59.0, 53.5 (Cꢀ
1’OOCH3), 36.0, 24.6, 21.4, 21.1, 20.9, 20.8, 20.7, 20.5; HRꢀMALDIꢀToF/MS m/z calcd for
C42H49NNaO19S [M + Na]+ 926.2517, found 926.2574.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
1,3,4,6ꢀTetraꢀOꢀacetylꢀ2ꢀdeoxyꢀ2ꢀ(2,2,2ꢀtrichloroethoxycarbonylamino)ꢀα/βꢀDꢀ
glucopyranoside (11)
To a solution of Dꢀglucosamine hydrochloride (20 g, 92.8 mmol) in H2O (250 mL) satd aq
NaHCO3 (250 mL) and 2,2,2ꢀtrichloroethyl chloroformate (14.05 mL, 102 mmol) were added and
stirred vigorously for 18 h at room temperature. The reaction mixture was filtered through a
fritted funnel and the white solid was dried under vacuum overnight. A solution of the solid in
pyridine (100 mL) was cooled to 0°C, to which Ac2O (100 mL) was added slowly, followed by
DMAP (500 mg, 41 mmol). The solution was warmed to room temperature and stirred for 12 h.
The solution was then diluted with CH2Cl2 (200 mL), washed with 1M HCl (3 x 100 mL), satd aq
NaHCO3 (2 x 100 mL), brine (100 mL), dried over Na2SO4, filtered, and concentrated in vacuo,
to afford glucosamine 11 as a white foam (44.3 g, 91%). The NMR spectra for 11 matched
those previously reported.52 1H NMR (400 MHz; CDCl3) δ: 6.22 (d, J = 3.7 Hz, 1H), 5.38 –
5.06 (m, 4H), 4.87 – 4.68 (m, 2H), 4.61 (d, J = 12.0 Hz, 1H), 4.32 – 3.88 (m, 6H), 2.18 –
2.00 (m, 13H); 13C NMR (100.5 Hz, CDCl3) δ: 171.2, 170.6, 169.1, 168.5, 154.0, 95.2,
90.4, 90.34, 74.7, 74.6, 70.4, 61.5, 53.2, 20.9, 20.8, 20.7, 20.6, 20.6, 20.5, 20.5; ESIꢀ
HRMS m/z calcd for C17H22Cl3NO11Na [M + Na]+ 544.0156, found 544.0168.
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 18 of 30
18
1
2
3
4
tertꢀButyldiphenylsilyl 3,4,6ꢀTetraꢀOꢀacetylꢀ2ꢀdeoxyꢀ2ꢀ(2,2,2ꢀtrichloroethoxycarbonylꢀ
5
6
amino)ꢀβꢀDꢀglucopyranoside (12)
7
8
9
A solution of 11 (10.3 g, 19.8 mmol) and hydrazinium acetate (1.9 g, 20.6 mmol) in DMF
(70 mL) was stirred for 20 min at room temperature. Then the reaction mixture was diluted with
EtOAc (400 mL), washed with saturated aqueous NaHCO3, brine, dried over Na2SO4, filtered,
and concentrated in vacuo. TBDPSCl (8.1 mL, 31.1 mmol) followed by imidazole (2.1 gm, 31.1
mmol) were added to the stirring solution of the crude residue in DMF (50 mL) at room
temperature. After 12 h, the reaction mixture was concentrated and diluted in EtOAc (200 mL),
washed with saturated aqueous NaHCO3, brine, dried over Na2SO4, filtered, and concentrated
in vacuo. The crude residue was purified by column chromatography over silica gel using 20%
EtOAc in hexanes to obtain 12 as a white foam (11.1 gm, 78%). The NMR spectra for 12
matched those previously reported.53 1H NMR (400 MHz; CDCl3) δ: 7.66 (m, 3H), 7.49 – 7.22
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
(m, 5H), 5.23 (d, J = 9.6 Hz, 1H, Hꢀ1), 5.13 – 4.90 (m, 2H, Hꢀ5, Hꢀ3), 4.75 (d, J = 12.0
Hz, 1H, 6ꢀCH2), 4.57 (m, 2H, 6ꢀCH2, Hꢀ4) , 4.06 (dd, J = 12.1, 5.7 Hz, 1H, 6ꢀCH2), 3.99
– 3.79 (m, 2H, Hꢀ2,Troc CH2), 3.40 (m, 1H, Troc CH2), 1.95 (m, 6H, 2CH3), 1.89 (s, 3H,
CH3), 1.06 (s, 9H, tBu); ; 13C NMR (100.5 Hz, CDCl3) δ: 170.8, 170.5, 169.4, 154.1, 135.9,
135.7, 135.7, 132.7, 132.6, 130.0, 129.9, 129.9, 127.6, 127.4, 96.0, 95.3, 74.6, 74.6,
72.4, 71.5, 62.2, 57.87, 26.7, 26.7, 20.6, 20.6, 20.6, 20.5, 20.5, 19.1; ESIꢀHRMS m/z
calcd for C31H38Cl3NO10SiNa [M + Na]+ 740.1228, found 740.1241.
tertꢀButyldiphenylsilyl 6ꢀOꢀbenzylꢀ2ꢀdeoxyꢀ2ꢀ(2,2,2ꢀtrichloroethoxycarbonylꢀamino)ꢀβꢀ
Dꢀ
glucopyranoside (14)
Trifluoroacetic acid (2.70 mL, 34.4 mmol) was added to a solution of 13 (4.7 gm, 6.9
mmol) and trimethylsilane (5.48 mL, 34.4 mmol) in CH2Cl2 (30 mL) at 0°C. The reaction mixture
was then warmed to room temperature and stirred for 3 h. The reaction mixture was
subsequently diluted in chloroform (20 mL), washed with a saturated aqueous solution of
ACS Paragon Plus Environment

Page 19 of 30
The Journal of Organic Chemistry
19
1
2
3
4
5
6
7
8
9
NaHCO3 (2 x 20 mL), brine, dried over MgSO4, filtered and concentrated. Purification of the
crude residue was performed by column chromatography over silica gel using 45% EtOAc in
hexanes as eluent, which afforded diol 14 as a white foam (4.2 gm, 89%). [α]D24 ꢀ13.1 (c = 1.0,
CHCl3), Rf 0.40 (hexane/EtOAc, 2:3); 1H NMR (400 MHz; CDCl3) δ: 7.70 (d, J = 7.6 Hz, 2H),
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
7.65 (d, J = 7.6 Hz, 2H), 7.39 (t, J = 7.2 Hz, 2H), 7.36ꢀ7.28 (m, 6H), 7.25ꢀ7.22 (m, 3H), 4.98 (d, J
= 7.6 Hz, 1H, NHTroc), 4.70 (d, J = 11.6 Hz, 1H, TrocCH2), 4.64 (d, J = 12.0 Hz, 1H, PhCH2),
4.50 (d, J = 12.0 Hz, 1H, PhCH2), 4.46 (d, J = 7.6 Hz, 1H, Hꢀ1), 4.43 (d, J = 12.0 Hz, 1H,
TrocCH2), 3.60ꢀ3.54 (m, 4H, Hꢀ2, Hꢀ4, Hꢀ6a, Hꢀ6b), 3.41 (d, J = 10.4 Hz, 1H, Hꢀ3), 3.38 (d, J =
2.8 Hz, 1H, OH), 3.14 (d, J = 4.8 Hz, 1H, Hꢀ5), 3.11 (br s, 1H, OH), 1.08 (s, 9H, tBu); 13C NMR
(100.5 Hz, CDCl3) δ: 155.1, 137.8, 136.0, 135.9, 135.8, 132.9, 132.7, 130.1, 129.8, 128.4,
127.7, 127.5, 95.9 (Cꢀ1), 95.2 (TrocCl3), 75.1, 75.0, 74.9, 74.1 (Cꢀ5), 73.7 (Cꢀ3), 72.4 (Cꢀ4), 70.0
(Cꢀ6), 59.7 (Cꢀ2), 26.8, 19.1; HRꢀMALDIꢀToF/MS m/z calcd for C32H38Cl3NNaO7Si [M + Na]+
704.1381, found 704.1384.
tertꢀButyldiphenylsilyl (2,3,4ꢀtriꢀOꢀbenzylꢀαꢀLꢀfucopyranosylꢀαꢀ(1→3)]ꢀ6ꢀOꢀbenzylꢀ2ꢀ
deoxyꢀ2ꢀ(2,2,2ꢀtrichloroethoxycarbonylꢀamino)ꢀβꢀDꢀglucopyranoside (16)
Trichloroacetimidate 15 (2.6 gm, 4.5 mmol), glucosamine acceptor 13 (2.4 gm, 3.5
mmol), and freshly activated 4 Å molecular sieves (3.2 gm) were stirred in distilled CH2Cl2 (50
mL) for 1 h at ꢀ30°C. TMSOTf (276 ꢁL, 1.5 mmol) was added dropwise and the reaction mixture
stirred for 1 h at ꢀ20°C. Progress of the reaction was monitored by TLC. Upon consumption of
the acceptor, the reaction mixture was quenched with diisopropylethylamine (2 mL). The
reaction mixture was diluted with CH2Cl2 (100 mL), and filtered through a pad of Celite. The
filtrate was successively washed with saturated aqueous solution of NaHCO3 (50 mL), brine,
dried over MgSO4, filtered, and concentrated in vacuo. The crude residue was flushed through
silica gel using 20% ethyl acetate in hexanes and concentrated to obtain a yellowish foam (4.1
gm). The foamy residue (4.1 gm), Et3SiH (1.8 mL, 11.4 mmol), and pulverized 4 Å molecular
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 20 of 30
20
1
2
3
4
5
6
7
8
9
sieves (12 gm) were stirred in distilled CH2Cl2 (50 mL) at room temperature for 1 h and then
cooled to ꢀ78°C and stirred for another 30 min. TfOH (774 ꢁL, 10.1 mmol) was then added
slowly and stirred for 1 h.48 Upon completion of the reaction as monitored by TLC, Et3N (5 mL)
was added and stirred for another 5 min before warming the reaction mixture to room
temperature. The reaction mixture was then diluted with CH2Cl2 (100 mL), filtered through
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Celite, concentrated in vacuo. The crude residue was purified by column chromatography over
silica gel eluting with 25% ethyl acetate in hexanes to obtain 16 as a white foam (2.95 gm,
76%). [α]D24 ꢀ45.5 (c = 1.0, CHCl3), Rf 0.30 (hexane/EtOAc, 1:4). 1H NMR (400 MHz; CDCl3) δ:
7.76 (dd, J = 1.2, 7.8 Hz, 2H), 7.70 (dd, J = 1.2, 7.8 Hz, 2H), 7.44ꢀ7.26 (m, 26H), 4.97 (d, J =
11.2 Hz, 1H, PhCH2), 4.90 (d, J = 8.4 Hz, 1H, TrocNH), 4.84ꢀ4.70 (m, 5H, Hꢀ1’, 4 PhCH2), 4.63
(d, J = 11.6 Hz, 1H, PhCH2), 4.63 (d, J = 11.6 Hz, 1H, PhCH2), 4.58 (d, J = 8.0 Hz, 1H, Hꢀ1),
4.51 (d, J = 12.0 Hz, 1H, PhCH2), 4.45 (d, J = 12.0 Hz, 1H, PhCH2), 4.30 (d, J = 11.2 Hz, 1H,
PhCH2), 4.15ꢀ4.07 (m, 1H, Hꢀ5’), 4.05 (dd, J = 3.4 Hz, 1H, Hꢀ2’), 3.94 (dd, J = 2.80, 10.0 Hz, Hꢀ
3’), 3.67ꢀ3.59 (m, 4H, Hꢀ2, Hꢀ6a, Hꢀ6b, Hꢀ4’), 3.52 (t, J = 9.0 Hz, 1H, Hꢀ4), 3.25 (t, J = 8.4 Hz, 1H,
Hꢀ5), 3.12 (dt, J = 3.6,10.0 Hz, 1H, Hꢀ3), 1.14 (d, J = 6.4 Hz, 3H, Fuc CH3ꢀ6’), 1.10 (s, 9H, tBu);
13C NMR (100.5 Hz, CDCl3) δ: 154.1, 138.7, 138.5, 138.4, 136.1, 136.0, 133.2, 132.9, 129.9,
129.7, 128.5, 128.4, 128.3, 127.9, 127.7, 127.6, 127.4, 98.9 (Cꢀ1’), 95.8 (Cꢀ1), 95.4 (TrocꢀCCl3),
84.8, 78.9 (Cꢀ3’), 77.6, 76.4 (C), 74.9 (Cꢀ4’), 73.8 (Cꢀ2’), 73.5 (Cꢀ3), 73.4, 69.9 (Cꢀ4), 69.2 (Cꢀ
5’), 68.1 (Cꢀ6), 58.1 (Cꢀ6), 26.8 (tBuꢀCH3), 19.2 (Fuc CH3ꢀ6’), 16.6 (tBuꢀC); HRꢀMALDIꢀToF/MS
m/z calcd for C59H66Cl3NNaO11Si [M + Na]+ 1120.3368, found 1120.3359.
tertꢀButyldiphenylsilyl methylꢀ5ꢀacetamidoꢀ7,8,9ꢀtriꢀOꢀacetylꢀ5ꢀN,4ꢀOꢀcarbonylꢀ3,5ꢀ
dideoxyꢀDꢀglyceroꢀαꢀ
Dꢀgalactoꢀnonꢀ2ꢀulopyranosylonateꢀ(2→3)ꢀ2,6ꢀdiꢀOꢀacetylꢀ6ꢀOꢀ
benzylꢀβꢀ
Dꢀgalactothiopyranosylꢀβꢀ(1→4)[2,3,4ꢀtriꢀOꢀbenzylꢀαꢀLꢀfucopyranosylꢀαꢀ(1→3)]ꢀ6ꢀ
Oꢀbenzylꢀ2ꢀdeoxyꢀ2ꢀ(2,2,2ꢀtrichloroethoxycarbonylꢀamino)ꢀβꢀDꢀglucopyranoside (17)
ACS Paragon Plus Environment

Page 21 of 30
The Journal of Organic Chemistry
21
1
2
3
4
5
6
7
8
9
Disaccharide 9 (720 mg, 0.78 mmol), disaccharide 16 (1.05 gm, 0.96 mmol), and freshly
activated 4 Å molecular sieves (800 mg) were stirred in freshly distilled CH2Cl2 (15 mL) for 1 h at
room temperature. The reaction mixture was cooled to ꢀ40°C before adding NIS (511 mg, 1.8
mmol) and TfOH (8.9 ꢁL, 0.1 mmol). Upon consumption of the donor, as determined by TLC,
the reaction mixture was quenched with diisopropylethylamine (2 mL). The reaction mixture was
then diluted with CH2Cl2 (50 mL) and filtered through Celite. The filtrate was successively
washed with saturated aqueous solution of Na2S2O3 (20 mL), NaHCO3 (20 mL), brine, dried
over MgSO4, filtered and concentrated in vacuo. The crude residue was purified by
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
chromatography over silica gel using 50% EtOAc in hexanes to obtain sLeX 17 as a white foam
(1.23 gm 86%). [α]D24 ꢀ22.5 (c = 1.0, CHCl3), Rf 0.52 (hexane/EtOAc, 1:1); 1H NMR (400 MHz,
CDCl3) δ: 7.67 (d, J = 6.8 Hz, 2H), 7.60 (d, J = 6.4 Hz, 2H), 7.44ꢀ7.15 (m, 31H), 5.68 (dd, J =
2.0, 8.4 Hz, 1H, Hꢀ7’’’), 5.56ꢀ5.52 (m, 1H, Hꢀ8’’’), 5.27 (dd, J = 2.6, 7.8 Hz, 1H, NHTroc), 5.23 (d,
J = 3.6 Hz, 1H, Hꢀ1’), 5.03 (d, J = 3.2 Hz, 1H, Hꢀ4’’), 4.95 (d, J = 12.0 Hz, 1H, PhCH2), 4.92 (d, J
= 10.4 Hz, 1H, PhCH2), 4.87 (d, J = 10.8 Hz, 1H, Hꢀ1), 4.76 (d, J = 8.4 Hz, 1H, Hꢀ1’’), 4.79 (d, J
= 11.6 Hz, 1H, PhCH2), 4.72 (d, J = 11.6 Hz, 1H, PhCH2), 4.68ꢀ4.52 (m, 4H, PhCH2), 4.49 (dd, J
= 3.8, 6.6 Hz, 1H, Hꢀ3’’), 4.39ꢀ4.25 (m, 5H, PhCH2, Hꢀ9a’’’), 4.19 (dd, J = 2.0, 10.0 Hz, 1H, Hꢀ6’’’),
4.09 (dd, J = 3.4, 10.2 Hz, 1H, Hꢀ2’), 4.04ꢀ3.91 (m, 5H, Hꢀ9b’’’, hꢀ4’’’), 3.80 (s, 3H, CO2Me), 3.76
(dd, J = 6.0, 7.2 Hz, 1H), 3.66 (dd, J = 3.4, 11.4 Hz, 1H), 3.61ꢀ3.53 (m, 3H, Hꢀ6a, Hꢀ6a’’, Hꢀ5’’’),
3.46 (dd, J = 5.2, 9.2 Hz, 1H), 3.42 (d, J = 11.2 Hz, 1H, Hꢀ6b’’), 3.39 (d, J = 11.6 Hz, 1H, Hꢀ6b),
3.28 (t, J = 8.8 Hz, 1H), 3.06 (d, J = 8.0 Hz, 1H, Hꢀ3’), 2.85 (dd, J = 3.4, 11.8 Hz, 1H, Hꢀ3e’’’),
2.45 (s, 3H), 2.07 (s, 3H), 2.05 (s, 3H), 2.00 (s, 3H), 1.93 (s, 3H), 1.84 (t, J = 12.4 Hz, 1H, Hꢀ
3a’’’), 1.83 (s, 3H), 1.19 (d, J = 6.4 Hz, 3H, Meꢀ5’), 1.03 (s, 9H, tBu); 13C NMR (100.5 Hz, CDCl3)
δ: 171.4, 170.5, 170.2, 169.6, 169.2, 169.0, 167.1, 153.5, 139.0, 138.7, 138.6, 138.5, 137.8,
135.9, 135.8, 133.5, 132.9, 129.7, 129.5, 128.4, 128.3, 128.2, 128.1, 128.0, 127.7, 127.6, 127.5,
127.4, 127.3, 127.1, 127.0, 126.9, 99.3 (Cꢀ1’’), 97.7 (Cꢀ2’’’), 97.1 (Cꢀ1’), 95.2 (Cꢀ1), 94.8 (CCl3),
79.8, 77.9 (Cꢀ3’), 75.9 (Cꢀ2’), 75.3 (Cꢀ6’’’), 74.7, 74.4, 74.3, 74.1, 73.5, 73.2 (Cꢀ7’’’), 73.1, 73.0,
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 22 of 30
22
1
2
3
4
5
6
7
8
9
72.2, 71.8, 71.4, 70.3, 68.2 (Cꢀ8’’’), 68.0, 67.9, 67.3 (Cꢀ4’’), 66.4, 62.5 (Cꢀ9’’’), 61.1, 59.1 (Cꢀ2),
53.4 (Cꢀ1’’’OOCH3), 35.9 (Cꢀ3’’’), 29.5, 28.4, 24.5, 21.3, 20.9, 20.8, 20.7, 20.6, 19.1 (tBu), 16.7
(Meꢀ5’); HRꢀMALDIꢀTOF/MS m/z calcd for C95H109Cl3N2NaO30Si [M + Na]+ 1913.5798, found
1913.5811.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Methylꢀ5ꢀacetamidoꢀ7,8,9ꢀtriꢀOꢀacetylꢀ5ꢀN,4ꢀOꢀcarbonylꢀ3,5ꢀdideoxyꢀ
galactoꢀnonꢀ2ꢀulopyranosylonateꢀ(2→3)ꢀ2,6ꢀdiꢀOꢀacetylꢀ6ꢀOꢀbenzylꢀβꢀ
galactothiopyranosylꢀβꢀ(1→4)[2,3,4ꢀtriꢀOꢀbenzylꢀαꢀLꢀfucopyranosylꢀαꢀ(1→3)]ꢀ6ꢀOꢀbenzylꢀ
DꢀglyceroꢀαꢀDꢀ
Dꢀ
2ꢀdeoxyꢀ2ꢀ(2,2,2ꢀtrichloroethoxycarbonylꢀamino)ꢀβꢀDꢀglucopyranosideꢀ1ꢀ
trichloroacetimidiate (18)
sLeX tetrasaccharide 17 (1.2 gm, 0.63 mmol) was dissolved in THF (50 mL) and was
cooled to 0°C. TBAF solution in THF (1N, 660 ꢁL, 0.66 mmol) and acetic acid (25 ꢁL) were
added and stirred for 48 h at room temperature. Upon completion of the starting material, as
determined by TLC, the solvent was removed in vacuo, and the crude reaction mixture was
filtered through silica gel using EtOAc. The filtrate was concentrated and dried overnight in
vacuo. The solid residue was dissolved in CH2Cl2 (12 mL) and cooled to 0°C.
Trichloroacetonitrile (660 ꢁL, 3.3 mmol) and DBU (48 ꢁL, 0.3 mmol) were added dropwise to the
reaction mixture and stirred for 1 h at room temperature. Upon completion of the reaction, the
solvent was removed at 10°C and the crude was purified by column chromatography over silica
gel using 40% EtOAc in hexanes. Product 18 was obtained as a white foam (729 mg 64%).
[α]D24 ꢀ65.1 (c = 1.0, CHCl3), Rf 0.51 (hexane/EtOAc, 1:1); 1H NMR (400 MHz; CDCl3) δ: 8.63 (d,
J = 2.0 Hz, 1H, CCl3C=NH), 7.37ꢀ7.26 (m, 20H), 7.24ꢀ7.20 (m, 5H), 6.67 (d, J = 6.4 Hz, 1H),
6.56 (s, 1H), 5.72 (dd, J = 2.0, 9.2 Hz, 1H), 5.63 (dd, J = 5.2, 8.8 Hz, 1H), 5.30 (s, 1H), 5.02 (d,
J = 11.2 Hz, 1H), 5.00 (d, J = 9.6 Hz, 1H), 4.91 (d, J = 12.0 Hz, 1H), 4.90 (d, J = 12.0 Hz, 1H),
4.86 (d, J = 10.0 Hz, 1H), 4.83 (d, J = 8.8 Hz, 1H), 4.82 (d, J = 9.0 Hz, 1H), 4.67 (d, J = 5.2 Hz,
1H), 4.64 (d, J = 95.2 Hz, 1H), 4.50 (dd, J = 1.6, 12.0 Hz, 1H), 4.54 (d, J = 12.0 Hz, 1H), 4.49
ACS Paragon Plus Environment

Page 23 of 30
The Journal of Organic Chemistry
23
1
2
3
4
(dt, J = 1.6, 10.0 Hz, 1H), 4.38 (d, J = 12.0 Hz, 1H), 4.37(t, J = 11.2 Hz, 1H), 4.35 (d, J = 12.8
5
6
7
8
Hz, 1H), 4.28 (d, J = 11.6 Hz, 1H), 4.24ꢀ4.14 (m, 3H), 4.12ꢀ3.95 (m, 5H), 3.92ꢀ3.85 (m, 3H), 3.82
(s, 1H), 3.81 (s, 3H, COOMe), 3.76 (d, J = 6.4 Hz, 1H), 3.61ꢀ3.56 (m, 2H), 3.46 (dd, J = 5.0, 9.4
Hz, 1H), 3.33 (t, J = 8.8 Hz, 1H), 2.88 (dd, J = 3.0, 11.8 Hz, 1H, Hꢀ3e’’’), 2.46 (s, 3H), 2.12 (s,
3H), 2.09 (s, 3H), 2.01 (s, 3H), 1.99 (s, 3H), 1.94 (s, 3H), 1.86 (t, J = 12.4 Hz, 1H, Hꢀ3a’’’), 1.15
(d, J = 6.4 Hz, 3H, Meꢀ5’); 13C NMR (100.5 Hz, CDCl3) δ: 171.4, 170.5, 170.3, 169.9, 169.2,
169.1, 160.1, 154.4, 153.5, 138.8, 138.7, 138.4, 138.0, 137.8, 128.5, 128.4, 128.4, 128.2, 128.1,
128.0, 127.7, 127.5, 127.4, 127.3, 127.2, 127.1, 99.6, 98.6, 97.7, 95.5, 94.3, 91.1, 79.6, 77.9,
75.9, 74.8, 74.6, 74.5, 74.2, 74.1, 73.9, 73.3, 73.1, 73.0, 72.3, 71.8, 71.7, 70.6, 68.1, 68.0, 67.9,
67.3, 67.3, 62.7, 59.1, 55.0, 53.4 (Cꢀ1’’’OOCH3), 35.9, 24.5, 21.3, 21.0, 20.9, 20.8, 20.7, 16.7
(CH3ꢀ5’); HRꢀMALDIꢀTOF/MS m/z calcd for C79H91Cl3N2NaO30 [(M ꢀ C2Cl3N) + Na]+ 1675.4620,
found 1675.4663.
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Methylꢀ5ꢀacetamidoꢀ7,8,9ꢀtriꢀOꢀacetylꢀ5ꢀN,4ꢀOꢀcarbonylꢀ3,5ꢀdideoxyꢀ
galactoꢀnonꢀ2ꢀulopyranosylonateꢀ(2→3)ꢀ2,6ꢀdiꢀOꢀacetylꢀ6ꢀOꢀbenzylꢀβꢀ
galactothiopyranosylꢀβꢀ(1→4)[2,3,4ꢀtriꢀOꢀbenzylꢀαꢀLꢀfucopyranosylꢀαꢀ(1→3)]ꢀ6ꢀOꢀbenzylꢀ
2ꢀdeoxyꢀ2ꢀ(2,2,2ꢀtrichloroethoxycarbonylꢀamino)ꢀβꢀ ꢀ1ꢀthioꢀglucopyranoside (19)
DꢀglyceroꢀαꢀDꢀ
Dꢀ
D
sLeX imidate 18 (718 mg, 0.4 mmol), thiophenol (82 ꢁL, 0.8 mmol), and 4 Å molecular
sieves (900 mg) were dissolved in CH2Cl2 (20 mL) and stirred for 1 h at room temperature
before cooling to ꢀ78°C. TMSꢀOTf (14.4 ꢁL, 0.08 mmol) was added slowly and the reaction
mixture was warmed to ꢀ50°C and stirred for 1 h. The reaction was then quenched by the
addition of triethylamine (1 mL) and warmed to room temperature. The mixture was then diluted
with CH2Cl2 (50 mL) and filtered through a pad of Celite. The filtrate was successively washed
with saturated aqueous solution of NaHCO3 (20 mL), brine, dried over MgSO4, filtered, and
concentrated in vacuo. The crude was purified by column chromatography over silica gel using
40% EtOAc in hexanes to obtain 19 as a white foam (572 mg 82%). [α]D24 ꢀ68.8 (c = 1.0,
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 24 of 30
24
1
2
3
4
CHCl3), Rf 0.49 (hexane/EtOAc, 1:1); 1H NMR (400 MHz, CDCl3) δ: 7.45 (dd, J = 1.2, 7.2 Hz,
5
6
7
8
2H), 7.39 (dd, J = 1.2, 6.8 Hz, 2H), 7.35ꢀ7.27 (m, 18H), 7.24ꢀ7.17 (m, 8H), 5.68 (dd, J = 1.8, 8.6
Hz, 1H), 5.66 (dd, J = 2.4, 12.0 Hz, 1H), 5.62 (dd, J = 2.0, 8.0 Hz, 1H), 5.21 (d, J = 3.2 Hz, 1H),
5.09ꢀ5.03 (m, 2H), 4.99 (dd, J = 8.4, 9.6 Hz, 1H), 4.96 (d, J = 11.6 Hz, 1H), 4.82 (d, J = 10.8 Hz,
1H), 4.79 (d, J = 10.0 Hz, 1H), 4.76 (d, J = 12.0 Hz, 1H), 4.73 (d, J = 12.0 Hz, 1H), 4.72 (d, J =
12.0 Hz, 1H), 4.68 (d, J = 12.4 Hz, 1H), 4.64 (d, J = 11.6 Hz, 1H), 4.60 (d, J = 12.0 Hz, 1H),
4.58 (d, J = 6.0 Hz, 1H), 4.56 (dd, J = 2.4, 10.0 Hz, 1H), 4.48 (d, J = 12.0 Hz, 1H), 4.43ꢀ4.35 (m,
3H), 4.29 (d, J = 12.0 Hz, 1H), 4.22 (dd, J = 2.2, 9.4 Hz, 1H), 4.16 (t, J = 7.8 Hz, 1H), 4.09 (dd, J
= 3.4, 10.2 Hz, 1H), 4.03 (dd, J = 6.0, 12.0 Hz, 1H), 3.97 (t, J = 8.0 Hz, 1H), 3.93 (dd, J = 3.2,
12.4 Hz, 1H), 3.89 (dd, J = 2.6, 11.8 Hz, 1H), 3.87ꢀ3.82 (m, 3H), 3.82 (s, 3H), 3.80 (t, J = 7.0 Hz,
1H), 3.66ꢀ3.60 (m, 2H), 3.57 (br s, 1H), 3.47 (dd, J = 5.4, 9.4 Hz, 1H), 3.30 (t, J = 8.8 Hz, 1H),
2.90 (dd, J = 3.0, 7.8 Hz, 1H), 2.46 (s, 3H), 2.11 (s, 3H), 2.10 (s, 3H), 2.02 (s, 3H), 1.98 (s, 3H),
1.89 (s, 3H), 1.86 (t, J = 11.8 Hz, 1H), 1.18 (d, J = 6.0 Hz, 3H); 13C NMR (100.5 Hz, CDCl3) δ:
171.5, 170.5, 170.4, 169.7, 169.4, 169.3, 167.2, 153.7, 153.5, 138.5, 138.7, 138.5, 138.4, 137.8,
132.5, 128.8, 128.5, 128.4, 128.3, 128.2, 127.8, 127.7, 127.6, 127.5, 127.4, 127.3, 127.2, 127.1,
99.4, 97.7, 97.5, 95.5, 84.7 (Cꢀ1), 79.9, 79.3, 77.5, 75.9, 74.5, 74.4, 74.3, 74.0, 73.9, 73.2, 73.1,
73.0, 72.9, 72.0, 71.4, 68.8, 68.2, 67.9, 67.2, 66.7, 62.8, 59.1, 56.9, 53.4 (Cꢀ1’’’OOCH3), 35.9,
24.6, 21.4, 20.9, 20.7, 20.6, 16.7; HRꢀMALDIꢀTOF/MS m/z calcd for C85H95Cl3N2NaO29S [M +
Na]+ 1767.4704, found 1767.4704.
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
ACS Paragon Plus Environment

Page 25 of 30
The Journal of Organic Chemistry
25
1
2
3
4
Nαꢀ(Fluorenꢀ9ꢀylmethoxycarbonyl)ꢀOꢀ(Methylꢀ5ꢀacetamidoꢀ7,8,9ꢀtriꢀOꢀacetylꢀ5ꢀN,4ꢀOꢀ
5
6
7
8
carbonylꢀ3,5ꢀdideoxyꢀ
acetylꢀ6ꢀOꢀbenzylꢀβꢀ ꢀgalactopyranosyl)ꢀ(1→4)ꢀOꢀ[(2,3,4ꢀtriꢀOꢀbenzylꢀαꢀLꢀfucopyranosyl)ꢀ
(1→3)ꢀ6ꢀOꢀbenzylꢀ2ꢀdeoxyꢀ1ꢀthioꢀ2ꢀ(2,2,2ꢀtrichloroethoxycarbonylꢀamino)ꢀβꢀ
glucopyranosyl]ꢀ(1→6)ꢀ[(2,3,4,6ꢀtetraꢀOꢀacetylꢀβꢀ ꢀgalactopyranosyl)ꢀ(1→3)ꢀOꢀ2ꢀazidoꢀ2ꢀ
deoxyꢀαꢀ ꢀgalactopyranosyl]ꢀLꢀthreonine tertꢀbutyl ester (1)
DꢀglyceroꢀαꢀDꢀgalactoꢀnonꢀ2ꢀulopyranosylonateꢀ(2→3)ꢀOꢀ(2,4ꢀdiꢀOꢀ
D
9
Dꢀ
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
D
D
Method A: sLeX thioglycoside 19 (523 mg, 0.30 mmol), Coreꢀ1 diol 20 (373 mg, 0.41
mmol), and freshly activated 4 Å molecular sieves (600 mg) were stirred in freshly distilled
CH2Cl2 (8 mL) for 1 h at room temperature. The reaction mixture was cooled to ꢀ60°C before
adding BSP (79 mg, 0.37 mmol) and Tf2O (57 ꢁL, 0.33 mmol). Upon consumption of the donor,
as determined by TLC, the reaction mixture was quenched with diisopropylethylamine (2 mL).
The reaction mixture was then diluted with CH2Cl2 (50 mL) and filtered through a pad of Celite.
The filtrate was successively washed with saturated aqueous solution of NaHCO3 (20 mL),
brine, dried over MgSO4, filtered, and concentrated in vacuo. The crude residue was purified by
chromatography over silica gel with 60% EtOAc in hexanes to obtain hexasaccharide 1 as a
white foam (619 mg, 81%).
Method B: sLeX thioglycoside 19 (506 mg, 0.29 mmol), Coreꢀ1 diol 20 (365 mg, 0.40
mmol), and freshly activated 4 Å molecular sieves (400 mg) were stirred in freshly distilled
CH2Cl2 (8 mL) for 1 h at room temperature. The reaction mixture was cooled to ꢀ40°C before
adding NIS (137 mg, 0.61 mmol) and TfOH (4.4 ꢁL, 0.05 mmol). Upon consumption of the
donor, as determined by TLC, the reaction mixture was quenched with diisopropylethylamine (2
mL). The reaction mixture was then diluted with CH2Cl2 (50 mL) and filtered through a pad of
Celite. The filtrate was successively washed with saturated aqueous solution of Na2S2O3 (20
mL), NaHCO3 (20 mL), brine, dried over MgSO4, filtered, and concentrated in vacuo. The crude
residue was purified by chromatography over silica gel, eluting with 60% EtOAc in hexanes to
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 26 of 30
26
1
2
3
4
obtain hexasaccharide 1 as a white foam (621 mg 84%). [α]D24 ꢀ12.7 (c = 1.0, CHCl3), Rf 0.36
(hexane/EtOAc, 1:4) 1H NMR (600 MHz, CDCl3) δ: 7.79 (d, J = 7.4 Hz, 2H, Fmoc H), 7.66 (d, J
= 7.2 Hz, 2H, Fmoc H), 7.45ꢀ7.38 (m, 4H, Fmoc H), 7.35ꢀꢀ7.20 (m, 25H, 5 Bn), 5.72 (d, J = 7.8
Hz, 1H, NHTroc), 5.69 (dd, J = 2.0, 8.8 Hz, 1H, Hꢀ7’’’’’), 5.67 (d, J = 10.4 Hz, 1H, FmocNH),
5.64ꢀ5.61 (m, 1H, Hꢀ8’’’’’), 5.40 (d, J = 3.2 Hz, 1H, Hꢀ4’), 5.31 (dd, J = 8.2, 10.4 Hz, 1H, Hꢀ2’),
5.24 (br s, 1H, Hꢀ1’’’), 5.08ꢀ4.92 (m, 5H, Hꢀ1, Hꢀ2, Hꢀ3’, Hꢀ4’’’’, PhCH2), 4.82 (d, J = 10.2, 1H, Hꢀ
1’’), 4.80 (d, J = 7.8 Hz, 1H, Hꢀ1’’’’), 4.79 (d, J = 12.6 Hz, 1H, PhCH2) 4.85ꢀ4.77 (m, 3H, Hꢀ1’, Hꢀ
1’’, PhCH2), 4.76ꢀ4.60 (m, 10H, Hꢀ3’’’’, Hꢀ1’’’’, 4PhCH2 ), 4.53 (dd, J = 3.2, 9.8 Hz, 1H, Hꢀ6a),
4.50 (dd, J = 3.4, 5.2 Hz, 1H), 4.48 (br s, 1H), 4.42 (d, J = 12.2 Hz, 1H, FmocCH2), 4.42ꢀ4.26
(m, 7H, TrocCH2,Thr Hꢀ2, Hꢀ6, TrocCH2, Thr Hꢀ1, FmocCH), 4.22 (dd, J = 2.2, 9.5 Hz, 1H, Hꢀ
6’’’’’), 4.14 (d, J = 7.4 Hz, 1H), 4.12 (d, J = 5.4 Hz, 1H), 4.11ꢀ4.09 (m, 2H), 4.07 (dd, J = 6.0,
10.8 Hz, 1H), 4.05ꢀ3.87 (m, 8H), 3.83 (s, 3H), 3.82ꢀ3.72 (m, 4H, CH3ꢀ1’’’’’, Hꢀ6a), 3.69ꢀ3.64 (m,
1H), 3.61 (t, J = 10.6 Hz, 1H), 3.60 (s, 1H), 3.50 (dd, J = 4.4, 9.7 Hz, 1H), 3.42ꢀ3.35 (m, 1H),
3.32 (t, J = 8.8 Hz, 1H), 2.90 (dd, J = 3.5, 11.8 Hz, 1H, Hꢀ3e’’’’’), 2.48 (s, 3H), 2.19 (s, 3H), 2.16
(s, 3H), 2.13 (s, 3H), 2.09 (s, 3H), 2.05 (s, 3H), 2.02 (s, 3H), 2.01 (s, 3H), 1.99 (s, 3H), 1.90 (s,
3H), 1.88 (t, J = 12.0 Hz, 1H, Hꢀ3a), 1.53 (s, 9H, tBu), 1.29 (d, J = 6.4 Hz, 3H, ThrꢀCH3), 1.20 (d,
J = 6.4 Hz, 3H, CH3ꢀ5’’’); 13C NMR (150.5 Hz, CDCl3) δ: 171.6, 170.6, 170.5, 170.3, 170.2,
169.8, 169.7, 169.6, 169.4, 169.2, 169.2, 156.9, 153.7, 153.5, 143,9, 143.8, 141.3, 138.7, 138.6,
138.3, 137.7, 128.5, 128.3, 128.2, 127.8, 127.7, 127.6, 127.4, 127.1, 127.1, 125.3, 120.0, 100.6
(Cꢀ1’’’’), 100.2, (Cꢀ1’), 100.1 (Cꢀ1’’), 99.5 (Cꢀ1), 97.6 (Cꢀ1’’’), 97.3 (Cꢀ2’’’’’), 95.6 (TrocꢀCCl3),
83.0, 80.0 (Cꢀ6’’’’’), 77.5, 76.4 (Cꢀ5’’’’), 75.9, 75.2 (Cꢀ9’’’’’), 74.6 (Cꢀ3’’), 74.3 (Cꢀ5’’), 73.8 (Cꢀ5’),
73.1 (Cꢀ7’’’’’), 73.0, 72.9, 72.0, 71.8, 71.3 (Cꢀ3’), 71.0 (Cꢀ3’’’’), 70.8, 70.3, 69.1 (Cꢀ6), 68.8, 68.2
(Cꢀ2’’’’), 67.9, 67.7 (Cꢀ8’’’’’), 67.4 (Cꢀ6’), 67.1 (Cꢀ6’’’’), 66.8 (Cꢀ6’’), 62.8, 61.2, 59.3 (Cꢀ5’’’’’), 59.0
(Cꢀ2), 58.4 (Cꢀ2’’), 53.5 (Cꢀ1’’’’’), 47.1 (FmocCH), 35.9 (Cꢀ3’’’’’), 29.7, 28.1, 24.7, 22.8, 21.4,21.1,
21.0, 20.8, 20.7, 19.2 (ThrꢀCH3), 16.8 (Cꢀ6’’’); HRꢀMALDIꢀTOF/MS m/z calcd for
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
C122H143Cl3N6NaO47 [M + Na]+ 2571.7947, found 2571.7983.
ACS Paragon Plus Environment

Page 27 of 30
The Journal of Organic Chemistry
27
1
2
3
4
Acknowledgments
5
6
7
8
We acknowledge support from the National Institute of Health (R01DK107405, U01GM116196,
RO1HL128237, P41GM103694).
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Supporting Information Available
Copies of 1H, 13C, and 2D NMR spectra. This material is available free of charge via the Internet
at http://pubs.acs.org.
References
(1)
(2)
(3)
Kolaczkowska, E.; Kubes, P. Nat. Rev. Immunol. 2013, 13, 159ꢀ175.
Schmidt, S.; Moser, M.; Sperandio, M. Mol. Immunol. 2013, 55, 49ꢀ58.
Wedepohl, S.; BecerenꢀBraun, F.; Riese, S.; Buscher, K.; Enders, S.; Bernhard, G.;
Kilian, K.; Blanchard, V.; Dernedde, J.; Tauber, R. Eur. J. Cell Biol. 2012, 91, 257ꢀ264.
Angiari, S. Autoimmun Rev 2015, 14, 984ꢀ995.
(4)
(5)
(6)
(7)
(8)
(9)
Zarbock, A.; Ley, K.; McEver, R. P.; Hidalgo, A. Blood 2011, 118, 6743ꢀ6751.
Mehta, P.; Cummings, R. D.; McEver, R. P. J. Biol. Chem. 1998, 273, 32506ꢀ32513.
McEver, R. P.; Cummings, R. D. J. Clin. Invest. 1997, 100, 485ꢀ491.
Girard, J.ꢀP.; Moussion, C.; Förster, R. Nat. Rev. Immunol. 2012, 12, 762ꢀ773.
Patel, M. S.; MirandaꢀNieves, D.; Chen, J.; Haller, C. A.; Chaikof, E. L. Transl Res 2017,
183, 1ꢀ13.
(10) Baert, F.; Noman, M.; Vermeire, S.; Van Assche, G.; D'Haens, G.; Carbonez, A.;
Rutgeerts, P. N. Engl. J. Med. 2003, 348, 601ꢀ608.
(11) Chang, J.; Patton, J. T.; Sarkar, A.; Ernst, B.; Magnani, J. L.; Frenette, P. S. Blood 2010,
116, 1779ꢀ1786.
(12) Preston, R. C.; Jakob, R. P.; Binder, F. P.; Sager, C. P.; Ernst, B.; Maier, T. J Mol Cell
Biol 2015, 8, 62ꢀ72.
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 28 of 30
28
1
2
3
4
5
6
7
8
(13) Bedard, P. W.; Clerin, V.; Sushkova, N.; Tchernychev, B.; Antrilli, T.; Resmini, C.; Keith,
J. C.; Hennan, J. K.; Kaila, N.; DeBernardo, S. J. Pharmacol. Exp. Ther. 2008, 324, 497ꢀ
506.
9
(14) Kaila, N.; Janz, K.; Huang, A.; Moretto, A.; DeBernardo, S.; Bedard, P. W.; Tam, S.;
Clerin, V.; Keith, J. C.; Tsao, D. H. H.; Sushkova, N.; Shaw, G. D.; Camphausen, R. T.;
Schaub, R. G.; Wang, Q. J. Med. Chem. 2007, 50, 40ꢀ64.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
(15) Davenpeck, K. L.; Berens, K. L.; Dixon, R. A.; Dupre, B.; Bochner, B. S. J. Allergy Clin.
Immunol. 2000, 105, 769ꢀ775.
(16) Leppänen, A.; White, S. P.; Helin, J.; McEver, R. P.; Cummings, R. D. J. Biol. Chem.
2000, 275, 39569ꢀ39578.
(17) Leppänen, A.; Parviainen, V.; AholaꢀIivarinen, E.; Kalkkinen, N.; Cummings, R. D.
Glycobiology 2010, 20, 1170ꢀ1185.
(18) Krishnamurthy, V. R.; Sardar, M. Y.; Yu, Y.; Song, X.; Haller, C.; Dai, E.; Wang, X.;
HanjayaꢀPutra, D.; Sun, L.; Morikis, V. Nat Commun 2015, 6, 6387.
(19) Nicolaou, K.; Hummel, C.; Iwabuchi, Y. J. Am. Chem. Soc. 1992, 114, 3126ꢀ3128.
(20) Akçay, G.; Ramphal, J. Y.; d’Alarcao, M.; Kumar, K. Tetrahedron Lett. 2015, 56, 109ꢀ
114.
(21) Lu, D.; Hu, Y.; He, X.; Sollogoub, M.; Zhang, Y. Carbohydr. Res. 2014, 383, 89ꢀ96.
(22) Vohra, Y.; Buskas, T.; Boons, G.ꢀJ. J. Org. Chem. 2009, 74, 6064ꢀ6071.
(23) Hanashima, S.; Castagner, B.; Esposito, D.; Nokami, T.; Seeberger, P. H. Org. Lett.
2007, 9, 1777ꢀ1779.
(24) Thoma, G.; Magnani, J. L.; Patton, J. T. Bioorg. Med. Chem. Lett. 2001, 11, 923ꢀ925.
(25) Gege, C.; Vogel, J.; Bendas, G.; Rothe, U.; Schmidt, R. R. Chem. Eur. J. 2000, 6, 111ꢀ
122.
(26) Misra, A. K.; Ding, Y.; Lowe, J. B.; Hindsgaul, O. Bioorg. Med. Chem. Lett. 2000, 10,
1505ꢀ1509.
ACS Paragon Plus Environment

Page 29 of 30
The Journal of Organic Chemistry
29
1
2
3
4
5
6
7
8
(27) Ellervik, U.; Magnusson, G. J. Org. Chem. 1998, 63, 9314ꢀ9322.
(28) Kiyoi, T.; Nakai, Y.; Kondo, H.; Ishida, H.; Kiso, M.; Hasegawa, A. Bioorg. Med. Chem.
1996, 4, 1167ꢀ1176.
9
(29) Danishefsky, S. J.; Gervay, J.; Peterson, J. M.; McDonald, F. E.; Koseki, K.; Griffith, D.
A.; Oriyama, T.; Marsden, S. P. J. Am. Chem. Soc. 1995, 117, 1940ꢀ1953.
(30) Kameyama, A.; Ishida, H.; Kiso, M.; Hasegawa, A. Carbohydr. Res. 1991, 209, C1ꢀC4.
(31) Kunz, H.; Unverzagt, C. Angew. Chem. Int. Ed. 1988, 27, 1697ꢀ1699.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
(32) Marqvorsen, M. H.; Pedersen, M. J.; Rasmussen, M. R.; Kristensen, S. K.; DahlꢀLassen,
R.; Jensen, H. H. J. Org. Chem. 2016, 82, 143ꢀ156.
(33) Crich, D.; Dudkin, V. J. Am. Chem. Soc. 2001, 123, 6819ꢀ6825.
(34) Liao, L.; Auzanneau, F.ꢀI. Org. Lett. 2003, 5, 2607ꢀ2610.
(35) Boons, G.ꢀJ.; Demchenko, A. V. Chem. Rev. 2000, 100, 4539ꢀ4566.
(36) Srivastava, V. K. Carbohydr. Res. 1982, 103, 286ꢀ292.
(37) Pertel, S. S.; Kononov, L. O.; Zinin, A. I.; Chirva, V. J.; Kakayan, E. S. Carbohydr. Res.
2012, 356, 172ꢀ179.
(38) Baumann, K.; Kowalczyk, D.; Kunz, H. Angew. Chem. Int. Ed. 2008, 47, 3445ꢀ3449.
(39) Baumann, K.; Kowalczyk, D.; Gutjahr, T.; Pieczyk, M.; Jones, C.; Wild, M. K.;
Vestweber, D.; Kunz, H. Angew. Chem. Int. Ed. 2009, 48, 3174ꢀ3178.
(40) Crich, D.; Li, W. J. Org. Chem. 2007, 72, 7794ꢀ7797.
(41) Crich, D.; Li, W. J. Org. Chem. 2007, 72, 2387ꢀ2391.
(42) Hsu, C. H.; Chu, K. C.; Lin, Y. S.; Han, J. L.; Peng, Y. S.; Ren, C. T.; Wu, C. Y.; Wong,
C. H. Chem. Eur. J. 2010, 16, 1754ꢀ1760.
(43) Ellervik, U.; Grundberg, H.; Magnusson, G. J. Org. Chem. 1998, 63, 9323ꢀ9338.
(44) Crich, D.; Li, W. J. Org. Chem. 2007, 72, 2387ꢀ2391.
(45) Hori, H.; Nakajima, T.; Nishida, Y.; Ohrui, H.; Meguro, H. Tetrahedron Lett. 1988, 29,
6317ꢀ6320.
ACS Paragon Plus Environment