Synthesis of lactosamine-based building blocks on a practical scale and investigations of their assembly for the preparation of19F-labelled LacNAc oligomers

Article abstract of DOI:10.1039/c8ob03066a

The ubiquitous disaccharide N-acetyllactosamine (LacNAc type 2, Galβ1,4GlcNAc) is often over-expressed on the surface of cancer cells where it is bound by tumour secreted galectins contributing to cancer-related processes such as metastasis, adhesion, tumour survival, and immune escape. To facilitate NMR investigations into the binding interactions between oligo-LacNAc structures and galectins, which can show both exo- and endo-binding behaviour, a library of regioselectively ¹⁹F-labelled oligo-LacNAc structures was required. Herein, the synthesis on a practical scale of various N-protected (Troc, Phth, TFAc) lactosamine donors is reported starting from commercially available lactosamine hydrochloride. Investigations into their glycosylations with lactosamine acceptors to form ¹⁹F-containing LacNAc oligomers showed that benzylated acceptors significantly improved the yields over acetylated ones, and that, gratifyingly, the almost untried N-trifluoroacetamide (NTFAc) protected donors, already containing the desired ¹⁹F-label, were found to be optimal, both considering reaction yields and purification of the glycosylation reactions. The NTFAc group of reducing end acceptors was introduced through N-amide transacylation of linker-equipped LacNAc structures. A [2 + 2] synthetic approach was optimized for the preparation of tetrasaccharide LacNAc/TFAc-dimers and also further expanded to the synthesis of hexasaccharide LacNAc/TFAc-trimer structures.

Full text of DOI:10.1039/c8ob03066a

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DOI: 10.1039/C8OB03066A
Organic & Biomolecular Chemistry
ARTICLE
Synthesis of lactosamine-based building blocks on a practical scale
19
and investigations of their assembly for the preparation of F-
labelled LacNAc oligomers
Received 00th January 20xx,
Accepted 00th January 20xx
Cecilia Romanò † and Stefan Oscarson*a
a
DOI: 10.1039/x0xx00000x
The ubiquitous disaccharide N-acetyllactosamine (LacNAc type 2, Galβ1,4GlcNAc) is often over-expressed on the surface of
cancer cells where it is bound by tumour secreted galectins contributing to cancer-related processes such as metastasis,
adhesion, tumour survival, and immune escape. To facilitate NMR investigations into the binding interactions between oligo-
LacNAc structures and galectins, which can show both exo- and endo-binding behaviour, a library of regioselectively 19F-
labelled oligo-LacNAc structures was required. Herein, the synthesis on a practical scale of various N-protected (Troc, Phth,
TFAc) lactosamine donors is reported starting from commercially available lactosamine hydrochloride. Investigations into
their glycosylations with lactosamine acceptors to form 19F-containing LacNAc oligomers showed that benzylated acceptors
significantly improved the yields over acetylated ones, and that, gratifyingly, the almost untried N-trifluoroacetamide
www.rsc.org/
(
NTFAc) protected donors, already containing the desired 19F-label, were found to be optimal, both considering reaction
yields and purification of the glycosylation reactions. The NTFAc group of reducing end acceptors was introduced through
N-amide transacylation of linker-equipped LacNAc structures. A [2+2] synthetic approach was optimized for the preparation
of tetrasaccharide LacNAc/TFAc-dimers and also further expanded to the synthesis of hexasaccharide LacNAc/TFAc-trimer
structures.
dynamics changes15 not involving the classical contacting
residues of the carbohydrate recognition domain (CRD) may
play a significant role in the recognition event. In this context,
Introduction
Oligo-N-acetyllactosamine and poly-N-acetyllactosamine,
there is no full understanding of the factors that influence the
observed binding preferences and binding modes even for
simple N-acetyllactosamines, linear poly-N-acetyllactosamines,
and branched ones. Interestingly, depending on the assay set-
up, discrepant binding specificities have been proposed. For
example, galectin-1 was found to bind both terminal and
internal galactose residues of a poly-LacNAc chain (endo-
[
βGal(1

4)βGlcNAc(1

3)]
n
, are ubiquitous constituents of N-
1
and O-glycans decorating the surface of mammalian cells.
These structures are natural ligands for proteins of the galectin
family with the binding interaction modulating cell-cell
adhesion, inflammatory and immune responses, and a variety
of disease associated processes, e.g., tumour development and
metastasis. Although all galectins possess similar carbohydrate
binding specificities and conserved consensus sequences
investigations with various methods, such as crystallography
and thermodynamic binding data, hemagglutination assays,
frontal affinity chromatography (FAC), nuclear magnetic
resonance spectroscopy (NMR)
molecular dynamics simulations,
with glycan microarrays, have shown that the elongation of
the LacNAc type 2 (Galβ1,4GlcNAc) scaffold, the introduction of
charges, and its modes of presentation as linear, branched or
multivalent oligo-structures, significantly modify the binding
affinity of galectins to their N-acetyllactosamine-based ligands.
Recent reports have suggested that structural14 and/or
2
11
1
13
16
binding behaviour) in H and C NMR studies and FAC
6
experiments. On the opposite, ELISA-type assays with
3
immobilized neoglycoproteins carrying poly-LacNAc chains
have shown that galectin-1 does not independently bind to
internal LacNAc motifs (exo-binding lectin behaviour).17
Similarly, contrasting reports into the binding preferences of
LacNAc oligomers by galectin-9 N-terminal CRD have been
3–5
5
6
7
–10
commonly coupled with
and binding experiments
7,11,12
13
6
,18
presented.
Further contributing to the poor understanding
of the dynamics of glycans recognition by galectins is the lack of
libraries of oligo-N-acetyllactosamine structures and analogues
19
thereof.
Recently it was shown that the presence of a fluorine probe on
carbohydrates, both as deoxy-fluoro compounds2
0,21
or
x y
F
) containing derivatives,22–24 can
fluoroacetamide (−NHCOCH
be exploited to follow protein-carbohydrate binding events via
a.School of Chemistry, University College Dublin, Belfield, Dublin 4, Ireland.
†
Present address: Department of Chemistry, Technical University of Denmark,
Kemitorvet 207, Kgs. Lyngby, Denmark.
corresponding author
19
F NMR spectroscopy. To further investigate this technique in
*
the context of glycan-galectin interactions, we are interested in
the design and synthesis of suitable di- and trimeric N-
Electronic Supplementary Information (ESI) available: [details of any supplementary
information available should be included here]. See DOI: 10.1039/x0xx00000x
This journal is © The Royal Society of Chemistry 20xx
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ARTICLE
Journal Name
making lactosamine hydrochloride an available starting
View Article Online
DOI: 10.1039/C8OB03066A
material in oligosaccharide synthesis. Herein we describe the
preparation of a set of lactosamine building blocks, starting
from
lactosamine
hydrochloride,
including
novel
trifluoroacetamide-containing donors and acceptors, and their
application in the synthesis of trifluoroacetamide containing
Figure 1: Trifluoroacetamide containing LacNAc/LacNTFAc oligosaccharides 1−2
and 3−5.
LacNAc/LacNTFAc tetrasaccharides
1 and 2. Additionally, the
scope is expanded to the synthesis of precursors of the trimeric
trifluoroacetamide−containing LacNAc/LacNTFAc structures
3−5 (Figure 1).
acetyllactosamines as galectin substrates bearing
a
trifluoroacetamide group (TFAc) as the fluorine probe on
different lactosamine units (Figure 1). The target
oligosaccharides are equipped with a 2-azidoethyl spacer to
provide a site of attachment for multivalent presentation or,
possibly, glycoarray preparation, while the alternate pattern in
the disposition of the trifluoroacetamide provides a way to
Results and discussion
Synthesis of first generation building blocks from lactosamine
hydrochloride
19
identify single LacNAc units in F NMR, with no loss of the
1
0
binding interaction.
Aside from enzymatic and chemoenzymatic methodologies,
In order to explore the reactivity of different lactosamine
derived glycosyl donors and acceptors, 2,2,2-trichloroethoxy
2
5–
27
chemical synthesis of di-, tri-, and tetrameric N- carbamate
(NHTroc)
bearing
donors,
glycosyl
acetyllactosamine derivatives has been reported earlier by Alais trichloroacetimidate
8
and thioglycoside 9, trifluoroacetamide
2
8,29
30
and Veyrieres,
Srivastava and Hindsgaul, Nilsson and (NHTFAc) protected thioglycoside 10, phthalimide (NPhth)
Norberg, Buskas et al., Severov et al.,33 Pazynina et al.,
3
1
32
34
protected thioglycoside 12, and its corresponding 3’,4’-O-
Mong et al., and more recently by Li and co-workers.36 With isopropylidene derivative 15, were prepared as depicted in
the exception of Li and co-workers, most of these syntheses Scheme 1. While donors 12 are fully acetylated, compound
3
5
8
−
take advantage of a monosaccharide approach from which 15 allows for the continued elongation to trimeric structures.
suitable disaccharide blocks are formed and coupled to obtain The NHTFAc group is orthogonal to the NHTroc group but not to
oligo-N-acetyllactosamine chains. Most recently, the the NPhth group, so the latter donor type cannot be used with
semisynthesis of LacNAc-extended complex-type biantennary NHTFAc acceptors for the synthesis of targets 1−5.
37
oligosaccharides has been described by Maki et al.
Lactosamine and N-acetyllactosamine can be prepared by trichloroacetimidate donor
We recently reported the synthesis of NHTroc substituted
starting from lactosamine
Since the introduction of the 2,2,2-
trichlorethoxy carbonyl moiety on the lactosamine scaffold
and recently adapted by Li and co-workers. A more efficient followed by peracetylation (Ac O, pyridine) yielded almost
exclusively the more stable α-acetate (81% yield), the
8
38
10
azidonitration of
aminophosphonium intermediate as shown by Lafont et al.
D
-lactal hexaacetate
or from an hydrochloride.
39
36
2
4
0–
route is represented by the Heyns rearrangement of lactulose
6
44
which has been optimised and upscaled to 300 kg batches,42
Scheme 1: Reagents and conditions: (a) i. TrocCl, NaHCO
°C, 45 min; (d) EtSH, BF ·Et O, dry CH Cl , 0 °C, 1 h, (e) i. Zn, CH
anhydride, Et N, MeOH, 50 °C, 2 h, iii. Ac O, Py, RT, overnight; (g) EtSH, BF
,2-DMP, p-TsOH, acetone, RT, overnight; (j) Ac O, Py, RT, 4 h; (k) MeONa, MeOH, RT, 4 h; (l) 2,2-DMP, p-TsOH, 80 °C, overnight; (m) Ac
aq. AcOH, 40 °C, 3 h; (o) i. CH C(OCH , dry CH
3
, H
2
O, RT, overnight, ii. Ac
CN/AcOH, RT, 2 h, ii. TFAA, Et
·Et O, TMSOTf, dry CH
2
O, Py, RT, overnight; (b) AcOH-EDA, THF, RT, overnight; (c) Cl
N, dry CH Cl , 0 °C, 3 h; (f) i. MeONa, MeOH, RT, 15 min, ii. Phthalic
Cl , 0 °C RT, overnight; (h) MeONa, MeOH, pH = 8−9, RT, 7 h; (i)
O, Py, RT, overnight; (n) 80%
CN, 135 °C, 3 h, ii. 80% aq. AcOH, 40 °C, 2 h.
3 2 2
CCN, DBU, dry CH Cl ,
0
3
2
2
2
3
3
2
2
3
2
3
2
2
2

2
2
2
3
3
)
3
3
CN, p-TsOH, RT, 2 h, ii. 80% aq. AcOH, RT, 2 h, (p) i. TFAA, CH
3
2
| J. Name., 2012, 00, 1-3
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DOI: 10.1039/C8OB03066A
Organic & Biomolecular Chemistry
ARTICLE
Donor
Acceptor
Promoter
NIS/TfOH
NIS/TfOH
NIS/AgOTf
Temperature
−10 °C
−40 °C  RT
−40 °C  RT
Time
2 h
30 min
2 h
Product
Yield
1
2
3
15
15
15
20
20
20
Hydrolysis
Hydrolysis
Complex mixture
Hydrolysis and 3’,4’-O-
isopropylidene
cleavage
2
h
4
15
20
DMTST
0 °C  RT
DMTST/2,4,6-
TTBP
DMTST/2,4,6-
TTBP
o/n
o/n
5
6
15
15
20
21
0 °C  RT
0 °C  RT
23, 25
45%
51%
24
7
8
9
12
10
10
9
21
21
21
22
22
DMTST
0 °C  RT
0 °C  RT
0 °C
0 °C  RT
0 °C
3 h
2 h
2 h
3 h
2 h
26
-
27
28
28
30%
-
40%
20%
45-56%
NIS/TfOH
NIS/TMSOTf
DMTST
1
0
1
1
8
TMSOTf
Table 1: [2+2] glycosylations tested with acceptors 20
−
22 and donors
8−
10,
12
,
15.
Chart 1: [2+2] glycosylations products
continuing preparation of the corresponding thioglycoside OH, NHAc), and 22 (3’-OH, NHTFAc) were prepared as
donor
by treatment with EtSH and a Lewis acid promoter (e.g., previously reported (Scheme 1).10 Briefly, lactosamine
BF ·Et O or TMSOTf) proved difficult. For this reason, hydrochloride was peracetylated and converted into its
trichloroacetimidate donor was first prepared by selective oxazoline derivative prior to the introduction of the 2-
anomeric acetate removal ( , 74%) followed by treatment azidoethyl linker, yielding compound 16 in an overall 40% yield.
with trichloroacetonitrile and DBU 85%), and Deacetylation ( 17, 91%) and regioselective introduction of
subsequently converted to thioglycoside in an 87% yield. From the 3’,4’-O-isopropylidene gave derivative 18 in a 72% yield.
donor , trifluoroacetamide derivative 10 was readily obtained Acetylation ( 19, 74%) followed by cleavage of the 3’,4’-O-
in two steps by cleavage of the carbamate moiety with Zn/AcOH isopropylidene ring under acidic conditions afforded the
in CH CN, followed by reaction with trifluoroacetic anhydride corresponding diol ( 20, 90%). Subsequent 3’,4’-O-orthoester
and Et N in CH Cl 10, 87%). NPhth donors 12 and 15 were formation and rearrangement in acidic conditions provided
9
3
2
8

7
(

8
,

9
9

3

3
2
2
(
both straightforwardly synthesized from lactosamine acceptor 21 in a 98% yield over two steps. Secondary amide N-
hydrochloride; reaction with phthalic anhydride in basic transacylation performed with trifluoroacetic anhydride (TFAA)
conditions, followed by acetylation gave 11 as a separable α/β in CH
mixture (α/β = 3:7, 56%). Reaction of the β-anomer of 11 with trifluoroacetamide derivative (
EtSH and BF ·Et O afforded the corresponding thioglycoside 12 product formation in only five minutes described in the work of
3
CN, followed by mild acidic work-up, afforded the desired

22, 70%). Contrarily to the fast
3
2
4
5
in an 85% yield. Further simplifying the synthetic approach, the Rota et al., the average reaction time on disaccharide 21 was
same reaction was performed on the α/β mixture, requiring the three hours, which was not shortened by microwave heating.
addition of catalytic amounts of TMSOTf to promote the
conversion of the α-anomer, globally affording donor 12 in a Glycosylations
6
2% yield. Compound 12 was then deacetylated under Zemplén
Performing efficient glycosylation reactions using the
synthesised set of building blocks proved difficult (Table 1, Chart
conditions ( 13, 97%) and treated with 2,2-dimethoxypropane

and catalytic amounts of p-TsOH to give the corresponding 3’,4’-
O-isopropylidene protected derivative 14 as the major product.
1
). Glycosylations of NPhth donor 15 with acceptor 20
promoted with either NIS/TfOH or NIS/AgOTf, afforded
exclusively donor hydrolysis (Entries 1−3), while
,
1
13
H and C NMR confirmed the formation of the correct
dioxolane ring. A quaternary carbon peak appeared at 110.6
ppm corresponding to the formation of a 5-membered ring, and
a concurrent down-field shift of the H-3’ and H-4’ signals was
observed in H NMR. Acetylation of 14 gave the desired donor
1
dimethyl(methylthio)sulfonium triflate (DMTST) activation
resulted in the hydrolysis of both the thioglycoside and the
acetal moiety of 15 (Entry 4). However, by adding 2,4,6-tri-tert-
butyl pyrimidine (2,4,6-TTBP) as an acid scavenger,
tetrasaccharide 23 was isolated, even though as an inseparable
1
5 in a 94% yield, on a multi-gram scale. Starting from
compound 16, glycosyl acceptors 20 (3’,4’-O-diol, NHAc), 21 (3’-
2
:1 mixture with the β(1

4) product 25 (Entry 5). Employing
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the same glycosylation conditions with 4’−OAc acceptor 21
resulted in the formation of tetrasaccharide 24 in a 51% yield
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DOI: 10.1039/C8OB03066A
(
Entry 6). Nevertheless, as the addition of the acid scavenger
substantially slowed the rate of the reaction, thus increasing the
number of observed by-products, non-buffered
a
DMTST−promoted coupling of fully acetylated NPhth donor 12
with acceptor 21 was tested (Entry 7). Unfortunately, the
corresponding product 26 was only isolated in 30% yield.
Moving to the coupling of NHTFAc donor 10 with acceptor 21
no reaction was observed with NIS/TfOH as promoting system
Entry 8), while tetrasaccharide 27 was obtained in a 40% yield
,
(
after activation by NIS/TMSOTf (Entry 9). An even lower product
yield was observed in a DMTST−promoted glycosylation of
Scheme 2: Reagents and conditions (a) BnBr, NaH (60% dispersion in mineral oil),
dry DMF, −5 °C, 3 h; (b) 80% aq. AcOH, 60 °C, 2 h; (c) i. CH
3
C(OCH
3 3
) , p-TsOH,
CH CN, RT, 40 min, ii. 80% aq. AcOH, RT, 1 h; (d) TFAA, Et N, CH
3
3
3
CN, 135 °C, 2 h.
NHTroc protected thioglycoside donor
9
and acceptor 22
(28,
2
0%, Entry 10), while a TMSOTf−promoted coupling of
acceptors. Benzylation (NaH, BnBr) of 3’,4’-O-isopropylidene
precursor 18 was performed at a reaction temperature of −5 °C.
By controlling the reaction time and dryness of reagents, the N-
benzylation byproduct was largely reduced giving compound 29
in a 68% yield on a 2 gram scale. Subsequent 3’,4’-O-
trichloroacetimidate
formation of tetrasaccharide 28 in variable but acceptable
yields (Entry 11). The latter glycosylation was scaled-up and
tetrasaccharide
8 with acceptor 22 resulted in the
1 was obtained after deprotections, as
10
previously reported.
isopropylidene cleavage at 60 °C in 80% aq. AcOH (30, 92%),
followed by acid catalysed 3’,4’-O-orthoester formation and its
subsequent rearrangement under acidic conditions, afforded
Synthesis of second generation acceptors and glycosylations
Across all tested couplings, the substantial amounts of acceptor 31 in a 92% yield over two steps. Acetamide N-
recovered unreacted acceptor suggested that compounds transacylation was carried out at 135 °C with TFAA and in the
4
5
2
0−22 have low reactivity, not well-matched to the one of the presence of triethylamine as acid scavenger yielding the
designed donors. Thus, a new set of benzyl ether protected corresponding trifluoroacetamide acceptor (
32, 58%).
building blocks was synthesized (Scheme 2) and investigated as
Donor
9
Acceptor
31
Product
Yield
87%
1
2
12
31
85%
3
4
10
9
31
32
94%
78%
2 2
Table 2: [2+2] glycosylations with acceptors 31−32 and donors 9−10, 12. General reagents and conditions: NIS, TfOH, −20 °C, dry CH Cl , 1 h.
Scheme 3: Reagents and conditions (a) NaBrO
3 2 2 4 2 2
, Na S O , H O, AcOEt, RT, 6 h; (b) i. TBAF (1M in THF), THF, RT, 6 h, ii. Ac O, Py, RT, overnight; (c) MeONa, MeOH, pH = 9, RT, overnight;
(
d) NaBrO , Na , H O, AcOEt, RT, 6 h; (e) MeONa, MeOH, pH = 9, RT, 7 h.
3
2
S
2
O
4
2
4
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DOI: 10.1039/C8OB03066A
Glycosylations of the acceptors 31 and 32 with donors
resulted in high to excellent yields of exclusively β(1
linked tetrasaccharides 34 36 (Table 2). The promoting system
of choice was NIS/TfOH at −20 °C in CH Cl . It should be noted
9
,
10, and
1
2

3)-
−
2
2
that no oxazoline formation was detected for NHTFAc donor 10
and no elimination occurred in case of NPhth donor 12 when
the thioglycoside was activated with a stochiometric amount of
TfOH (0.8−1 equivalents). NHTFAc protected donors
46
(
glucosamine scaffold) were introduced early but have been
used less often than carbamate and NPhth protected donors.
Regarding NHTFAc donors based on the LacNAc scaffold, only
two reports could be found; a LacNAc type 1 (Gal1,3GlcNAc)
Scheme 4: Reagents and conditions (a) BnBr, TBAI, NaH (60% dispersion in
NHTFAc-protected thioglycoside donor employed in
a
mineral oil), dry DMF, 0 °C
TrocCl, NaHCO , THF, RT, 3 h; (c) i. H
CH Cl , 0 °C, 1 h.

RT, 3 h; (b) i. H
2 2 2
N-NH ·H O, EtOH, reflux, 6 h, ii.
3
2
N-NH ·H O, EtOH, reflux, 10 h, ii. TFAA, Py,
2
2
4
7
glycosylation with a lactose-derived acceptor, and, very
recently, a LacNAc type 2 NHTFAc-protected imidate donor that
2
2
4
8
was reacted with mannose-based acceptors. In both these
reports, the LacNTFAc was produced from monosaccharide
precursors and in neither report was a comparison made with
other differently N-protected donors. Indeed, earlier
assessments, considering glycosylation yields, between
differently N-protected donors have showed the NHTFAc
protected donors to be, at the best, equivalent to the other
yield. Final Zemplén deacetylation gave tetrasaccharide
5% yield. Deprotection of 35 proceeded similarly; benzyl
groups were first removed under oxidative conditions to give
compound 39 71%), followed by Zemplén deacetylation
affording compound in a 55% yield.
1 in a
6
(

2
Expanding the building block library and the strategic
assembly to hexasaccharides
4
9
donors. Here, however and most gratifyingly, the lactosamine
NHTFAc donor 10 afforded the highest yield and cleanest
reaction with benzylated acceptor 31 (Table 2, Entry 3), a
characteristic that is reflected on all glycosylations involving
NHTFAc protected donors reported in this work (see Table 3,
Entry 2 and Table 4, Entry 2). Tetrasaccharides 35 and 36 were
selected to study the deprotection of the newly synthesised
To extend the developed methodology to the preparation of
larger LacNAc oligomers, a new series of 3’,4’-O-isopropylidene
donors bearing benzyl ether protecting groups instead of acetyl
esters was synthesized in order to prevent low yielding [2+4]
couplings. Benzylations of the lactosamine scaffold in the
presence of the carbamate and NHTFAc as amine protecting
groups were deemed not feasible. However, benzylation of
NPhth derivatives is possible but care has to be taken to avoid
oligomers in order to allow for the synthesis of compounds
2
and , respectively, as depicted in Scheme 3. Importantly, both
1
protected tetrasaccharides 35 and 36 have already the desired
trifluoroacetamide probe installed, thus reducing the number
of steps necessary to afford the desired final compounds. Benzyl
groups on compound 36 were removed under oxidative
36
opening of the phthalimide ring. Compound 14 was
benzylated with BnBr and NaH in dry DMF giving 40 in a 54%
yield on a multi-gram scale (Scheme 4). Compound 40 was then
readily converted into its NHTroc protected derivative 41 and
NHTFAc analogue 42 by basic cleavage of the phthalimide
moiety and treatment with either 2,2,2-trichloroethyl
50
3 2 2 4
conditions by treatment with NaBrO /Na S O in AcOEt, thus
maintaining the azide functionality and yielding compound 37
in 73% yield. Removal of the Troc carbamate, again without
affecting the azide, was performed with TBAF in THF as
chloroformate in basic conditions (
trifluoroacetic anhydride in pyridine/CH

41, 63%) or with
2
Cl (42, 79%).
2
5
1
52
previously reported by Jacquemard et al. and Huang et al.,
followed by acetylation with Ac O in pyridine to give 38 in a 68%
2
Donor
40
Acceptor
31
Product
Time Yield
90 min 64%
1
2
3
42
41
31
32
20 min 81%
1 h
78%
2 2
Table 3: [2+2] glycosylations with donors 40−42.General reagents and conditions: NIS, TfOH, −25 °C, dry CH Cl , 1 h
This journal is © The Royal Society of Chemistry 20xx
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ARTICLE
Journal Name
View Article Online
DOI: 10.1039/C8OB03066A
Scheme 5: Reagents and conditions (a) i. TBAF (1M in THF), THF, RT, 8 h, ii. Ac
2
O, Py, RT, overnight; (b) 80% aq. AcOH, 70 °C, 4−5 h; (c) i. CH
3
C(OCH
3
)
3
, p-TsOH, CH
3
CN, RT, 30 min, ii.
80% aq. AcOH, RT, 1 h; (d) i. H
2
N-NH ·H
2 2
O, EtOH, reflux, 8 h, ii. Ac
2
O, Py, RT, overnight; (e) 80% aq. AcOH, 60 °C, 6 h; (f) i. CH
3
C(OCH
3
)
3
, p-TsOH, CH CN, RT, 30 min, ii. 80% aq. AcOH,
3
RT, 1 h.
Donor
9
Acceptor
50
Product
Yield
1
2
3
4
55%
10
9
55
51
52
80%
-
9
58%
The efficiency of the new set of donor building blocks was 58, but a complex mixture of compounds (Table 4, Entry 3).
tested in NIS/TfOH promoted glycosylations with acceptors 31 However, conversion of the Troc moiety of 45 to the
and 32. All three glycosylations were completely stereoselective corresponding acetamide (
-(1 3) linkage formation) with no 3’,4’-O-isopropylidene preparation of acceptor 52 with the same methodology (as
cleavage observed during the reaction time, and afforded the depicted in Scheme 5). Coupling of 52 with donor afforded
46, 78%) allowed the subsequent
(β

9
desired products in good yields (Table 3), but still lower hexasaccharide 59 in 58% yield (Table 4, Entry 4, compare Table
compared to the corresponding glycosylations with acetylated 1, Entry 1 (87%)).
donors (Table 2). Tetrasaccharides 43
into acceptors using the same reaction sequence as before;
’,4’-O-isopropylidene removal, 3’,4’-O-orthoester formation,
and its rearrangement in acidic conditions to afford 3’’’-OH
acceptors 50 51, and 55 (Scheme 5). For compound 43, an
additional step was performed to convert the phthalimide into
an acetamide moiety (
coupling of NHTroc thioglycoside
−45 were then converted
3
Experimental
General methods
,

53, 92%). The NIS/TfOH-promoted Unless noted, chemical reagents and solvents were used
with acceptor 50 gave without further purification from commercial sources.
9
hexasaccharide 56 in a 55% yield (Table 4, Entry 1), while Lactosamine hydrochloride was purchased from Glycom A/S.
NHTFAc donor 10 gave hexasaccharide 57 in an 80% yield, when Dry solvents as CH Cl , Et O, and THF were obtained from a
2
2
2
coupled with acceptor 55 (Table 4, Entry 2). Again, no oxazoline PureSolv-ENTM solvent purification system (Innovation
formation was observed with the NHTFAc-type donor 10.These Technology Inc). Concentration in vacuo was performed using a
yields can be compared with the corresponding [2+2] couplings Buchi rotary evaporator. The
in Table 2, Entries 4 (78%) and Entry 3 (94%), respectively. relative to TMS in CDCl ) were recorded with Varian
1
H/13C/19F NMR spectra (δ in ppm,
3
NIS/TfOH promoted glycosylation of NHTroc thioglycoside
with acceptor 51 did not afford the expected hexasaccharide 500/125ꢀMHz) at 25 °C. Assignments were aided by
9
spectrometers (Varian, Palo Alto, CA, USA) (400/101ꢀMHz or
H- H
1
1
6
| J. Name., 2012, 00, 1-3
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Journal Name
ARTICLE
1
13
and H- C correlation experiments. HRMS spectra were (C-5’), 74.4 (CH
2
Ph, CH
2 3
CCl ), 73.7 (CH
2
Ph), 73.6 (CH
2
Ph), 72.7 (C-5’’),
View Article Online
recorded on a micromass LCT instrument from Waters and 72.6 (C-5), 72.3 (C-3’’), 71.1 (C-3’’’), 70.9 (C-D5O’’’I):,1609.1.083(9C/C-48’O),B6093.036(6CA-
LaserToF LT3 Plus MALDI-TOF (DHAP Matrix). LRMS spectra 2’’’), 68.5 (OCH
were recorded on a Waters micromass Quattro Micro LC- 61.0 (C-6’’’), 57.2 (C-2), 56.4 (C-2’’), 50.7 (OCH
MS/MS instrument using electrospray ionisation (ESI) in either (NHCOCH ), 20.94, 20.89, 20.84, 20.80, 20.78, 20.76, 20.6 (7
3 5
positive or negative mode. Optical rotations were recorded on OCOCH ); HRMS (ESI ): m/z calcd for C73H88Cl N O29: 1626.4528
2
CH
2 3
N ), 68.2 (C-6, C-6’), 66.8 (C-4’’’), 61.57 (C-6’’),
2
CH ), 23.8
2 3
N
3
+
3
+
a Perkin-Elmer polarimeter (Model 343) at the sodium D-line [M+Na] ; found 1626.4451.
589ꢀnm) at 20 °C using a 1ꢀdm cell and are not corrected. Silica
gel chromatography was carried out using Davisil LC60A (Grace 2-Azidoethyl (2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl)-(14)-
tech., Columbia, MD, USA) SiO (40–63ꢀμm) silica gel. All (3,6-di-O-acetyl-2-deoxy-2-phthalimido-β-D-glucopyranosyl)-
reactions were monitored by thin-layer chromatography (TLC). (13)-(4-O-acetyl-2,6-di-O-benzyl-β-D-galactopyranosyl)-(14)-
TLC was performed on Merck DC-Alufolien plates precoated 2-acetamido-3,6-di-O-benzyl-2-deoxy-β-D-glucopyranoside (34) R
(
2
f
=
20
1
with silica gel 60 F254. They were visualised with UV-light 0.47, Tol/Acetone 7:3; [훼]D = +15.4 (c 1.0, CHCl
254ꢀnm) fluorescence quenching, and/or by charring with an CDCl ) δ 7.37 – 7.13 (m, 22H, HAr), 6.93 (dd, J = 6.6, 2.9 Hz, 2H, HAr),
% H SO dip and/or ninhydrin dip. Deprotected sugars were 5.77 (dd, J = 10.6, 8.7 Hz, 1H, H-3’’), 5.65 (d, J = 7.3 Hz, 1H, NHCOCH ),
3
); H NMR (500 MHz,
(
8
3
2
4
3
lyophilised using a freeze-dryer Alpha 1-2 Ldplus (Christ Ltd.), 5.53 (d, J = 8.2 Hz, 1H, H-1’’), 5.37 (ad, J = 3.5 Hz, 1H, H-4’), 5.33 (dd,
with a pressure of 0.035 mbar and ice condenser temperature J = 3.5, 1.2 Hz, 1H, H-4’’’), 5.13 (dd, J = 10.4, 7.9 Hz, 1H, H-2’’’), 4.97
−
55 °C.
(dd, J = 10.4, 3.5 Hz, 1H, H-3’’’), 4.93 (d, J = 7.6 Hz, 1H, H-1), 4.84 (d, J
10.9 Hz, 1H, CHHPh), 4.72 (dd, J = 11.9, 2.3 Hz, 1H, H-6’’a), 4.56 (d,
J = 7.9 Hz, 1H, H-1’’’), 4.50 (d, J = 12.2 Hz, 1H, CHHPh), 4.48 – 4.44 (m,
=
General procedure for [2+2] glycosylation for compounds 33−36
Acceptor (1 eq) and donor (1.2−1.3 eq) were dissolved in dry
CH Cl (0.018 M) together with 4Å molecular sieves. The
2 2
2
4
H, CHHPh, CHHPh), 4.35 – 4.22 (m, 4H, H-1’ CHHPh, CHHPh, CHHPh),
.13 (dd, J = 10.7, 8.4 Hz, 1H, H-2’’), 4.10 – 4.02 (m, 5H, H-3, CHHPh,
H-6’’’a, H-6’’’b, H-6’’b), 3.95 (ddd, J = 10.8, 5.2, 3.4 Hz, 1H,
OCHHCH ), 3.92 – 3.83 (m, 3H, H-5’’’, H-4, H-4’’), 3.74 (ddd, J =
0.0, 4.2, 2.4 Hz, 1H, H-5’’), 3.63 – 3.54 (m, 3H, OCHHCH , H-6’a,
H-3’), 3.51 (at, J = 6.2 Hz, 1H, H-5’), 3.45 – 3.23 (m, 6H, OCH CHHN
H-6’b, H-6a, H-6b, H-5, H-2’), 3.19 (ddd, J = 13.3, 5.2, 3.3 Hz, 1H,
OCH CHHN ), 3.11 (dd, J = 9.8, 7.6 Hz, 1H, H-2), 2.13 (s, 3H, OCOCH ),
mixture was stirred for 1 hour, then NIS (1.5 eq) was added and
the mixture was cooled to −20 °C. TfOH (0.5 eq) was then added
dropwise and the reaction was stirred for 30 minutes at the
same temperature, then quenched with Et N, filtered over
3
Celite and evaporated in vacuo. Crude was purified by flash
2 3
N
1
2 3
N
2
3
,
2
3
3
column chromatography (see Supplementary Information).
2
2
1
.09 (s, 3H, OCOCH
.00 (s, 3H, OCOCH
3
), 2.08 (s, 3H, OCOCH
), 1.97 (s, 3H, OCOCH
3
), 2.04 (s, 3H, OCOCH
), 1.87 (s, 3H, OCOCH
3
),
),
3
3
3
2
-Azidoethyl (2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl)-(14)-
.85 (s, 3H, NHCOCH
), 170.46, 170.3, 170.2, 170.1, 169.72, 169.66, 169.1 (7
), 138.9 (CAr), 138.1 (CAr), 138.0 (CAr), 137.9 (CAr), 129.0,
3
); 13C NMR (126 MHz, CDCl
3
) δ 170.52
[3,6-di-O-acetyl-2-deoxy-2-(2,2,2-trichloroethoxycarbonylamino)-
(
NHCOCH
3
β-D-glucopyranosyl]-(13)-(4-O-acetyl-2,6-di-O-benzyl-β-D-
galactopyranosyl)-(14)-2-acetamido-3,6-di-O-benzyl-2-deoxy-β-
OCOCH
3
2
0
128.3, 128.2, 128.1, 128.0, 127.8, 127.7, 127.6, 127.5, 127.1, 126.7,
25.3 (20 CAr), 102.1 (C-1’), 101.2 (C-1’’’), 99.3 (C-1), 98.1 (C-1’’), 78.6
C-2’), 78.5 (C-3’), 77.2 (C-3), 76.7, 76.4 (C-4’’, C-4), 74.8 (C-5), 74.4
CH Ph), 74.2 (CH Ph), 73.5 (CH Ph), 73.1 (CH Ph), 72.4 (C-5’’, C-5’),
1.0 (C-4’’’), 70.9 (C-3’’), 70.6 (C-5’’’), 70.1 (C-4’), 69.1 (C-2’’’), 68.3,
8.2, 67.7 (C-6’, C-6, OCH CH ), 66.6 (C-4’’’), 61.1 (C-6’’), 60.6 (C-
’’’), 57.1 (C-2), 55.3 (C-2’’), 50.6 (OCH
D-glucopyranoside (33) R
f
= 0.34, Tol/Acetone 7:3; [훼] = +2.6 (c 1.0,
D
1
(
1
CHCl
3
); H NMR (500 MHz, CDCl
3
) δ 7.44 – 7.20 (m, 20H, HAr), 5.72 (d,
), 5.38 – 5.33 (m, 2H, H-4’, H-4’’’), 5.10 (dd, J
10.4, 7.9 Hz, 1H, H-2’’’), 5.02 (d, J = 7.7 Hz, 1H, H-1), 4.96 (dd, J =
0.4, 3.4 Hz, 1H, H-3’’’), 4.92 (d, J = 10.9 Hz, 1H, CHHPh), 4.86 (d, J =
1.7 Hz, 1H, CHHPh), 4.77 – 4.72 (m, 1H, H-3’’), 4.67 (dd, J = 12.1,
J = 7.4 Hz, 1H, NHCOCH
3
2
2
2
2
=
1
1
1
7
6
6
2
C
2
2 3
N
2
CH
2
N
3
), 23.6 (NHCOCH
3
), 20.8,
0.2 Hz, 2H, CHHPh, CHHCCl
Ph, CHHCCl , H-6’’a, NHCOCH
H, CHHPh), 4.24 – 4.17 (m, 1H, H-3), 4.09 (d, J = 6.8 Hz, 2H, H-6a’’’,
3
), 4.61 – 4.38 (m, 11H, H-1’’, H-1’’’, H-1’,
+
0.74, 20.65, 20.6, 20.5 (7 OCOCH
3
); HRMS (ESI ): m/z calcd for
CH
2
3
2
CCl , CH Ph), 4.31 (d, J = 11.8 Hz,
3
2
+
78
H
89
5
N O
29: 1560.5721 [M+H] ; found: 1560.5714.
1
H-6b’’’), 4.06 – 3.97 (m, 3H, H-6a’’, OCHHCH
2
N
3
, H-4), 3.88 – 3.84 (m,
2
-Azidoethyl (2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl)-(14)-
1
1
H, H-5’’’), 3.80 (dd, J = 10.9, 3.8 Hz, 1H, H-6a), 3.74 (at, J = 9.3 Hz,
(3,6-di-O-acetyl-2-deoxy-2-trifluoroacetamido-β-D-
H, H-4’’), 3.69 – 3.63 (m, 3H, H-6b, OCHHCH
2
N
3
, H-3’), 3.61 – 3.54
, H-5’’, H-5’, H-2’),
.36 (adt, J = 6.9, 3.1 Hz, 2H, H-6a’, H-6b’), 3.27 – 3.17 (m, 2H,
CHHN , H-2), 2.14 (s, 3H, OCOCH ), 2.08 (s, 3H, OCOCH ), 2.06
s, 3H, OCOCH ), 2.05 (s, 3H, OCOCH ), 2.01 (s, 3H, OCOCH ), 1.99 (s,
), 1.97 (s, 3H, OCOCH ), 1.90 (s, 3H, NHCOCH
); 13C NMR
) δ 170.7 (NHCOCH ), 170.6, 170.5, 170.4, 170.23,
70.17, 170.0, 169.3 (7 OCOCH ), 154.2 (NHCO CH CCl ), 138.9,
glucopyranosyl)-(13)-(4-O-acetyl-2,6-di-O-benzyl-β-D-
(
2 3
m, 2H, H-2’’, H-5), 3.53 – 3.42 (m, 4H, OCH CHHN
galactopyranosyl)-(14)-2-acetamido-3,6-di-O-benzyl-2-deoxy-β-
3
OCH
(
3
2
0
D-glucopyranoside (35) R
CHCl
f
= 0.26, Tol/AcOEt 1:1; [훼] = -0.8 (c 1.0,
D
2
3
3
3
1
3
); H NMR (500 MHz, CDCl
3
): δ 7.53 – 7.10 (m, 20H, HAr), 6.22
), 5.74 (d, J = 7.4 Hz, 1H, NHCOCH ), 5.43
5.30 (m, 2H, H-4’, H-4’’’), 5.12 (dd, J = 10.5, 7.9 Hz, 1H, H-2’’’), 5.02
4.96 (m, 2H, H-1, H-3’’’), 4.92 – 4.86 (m, 2H, H-3’’, CHHPh), 4.82 (d,
3
3
3
(
d, J = 9.5 Hz, 1H, NHCOCF
3
3
H, OCOCH
3
3
3
–
–
(
126 MHz, CDCl
3
3
1
1
1
1
8
3
2
2
3
J = 11.8 Hz, 1H, CHHPh), 4.71 (d, J = 7.8 Hz, 1H, H-1’’), 4.68 – 4.62 (m,
H, CHHPh, H-6’’a), 4.57 (d, J = 11.0 Hz, 1H, CHHPh), 4.53 (d, J = 7.9
Hz, 1H, H-1’’’), 4.51 – 4.45 (m, 2H, CHHPh, CHHPh), 4.44 (d, J = 7.8 Hz,
H, H-1’), 4.40 (d, J = 12.1 Hz, 1H, CHHPh), 4.31 (d, J = 11.9 Hz, 1H,
38.4, 138.2, 138.1 (4 CAr), 129.2, 128.9, 128.6, 128.5, 128.31,
28.29, 128.2, 128.1, 128.0, 127.82, 127.80, 127.7, 127.1 (20 CAr),
2
02.4 (C-1’), 101.4 (C-1’’’), 101.1 (C-1’’), 99.6 (C-1), 95.7 (CH
2 3
CCl ),
1
0.7 (C-2’), 77.4 (H-3), 76.6 (C-3’, H-4), 76.3 (H-4’’), 75.2 (CH Ph), 75.1
2
CHHPh), 4.18 (dd, J = 9.6, 8.2 Hz, 1H, H-4), 4.14 – 4.08 (m, 2H, H-6’’’a,
This journal is © The Royal Society of Chemistry 20xx
J. Name., 2013, 00, 1-3 | 7
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ARTICLE
Journal Name
H-6’’’b), 4.06 (dd, J = 12.1, 4.4 Hz, 1H, H-6’’b), 4.04 – 3.95 (m, 2H, H- deoxy-2-trifluoroacetamido-β-D-glucopyranoside) (1)ViTewo AartiscoleluOtniloinne
2 3
N
), 3.98 – 3.91 (m, 1H, H-2’’), 3.91 – 3.86 (m, 1H, H-5’’’), of 36 (110 mg, 66 μmol) in AcOEt (880 μL) DwOaIs: 1a0d.1d0e3d9/aC8soOlBu0ti3o0n66oAf
3
3
3
, OCHHCH
.82 (at, J = 9.0 Hz, 1H, H-4’’), 3.77 (dd, J = 10.8, 3.9 Hz, 1H, H-6a), NaBrO
.73 – 3.62 (m, 3H, H-3’, OCHHCH , H-6b), 3.58 (at, J = 6.4 Hz, 1H, Na
CHHN
m, 2H, H-6’a, H-6’b), 3.24 (ddd, J = 14.8, 6.2, 3.1 Hz, 1H, with 10% Na
OCH CHHN ), 2.15 (s, 3H, OCOCH ), 2.08 (s, 3H, OCOCH ), 2.07 (s, 3H, and evaporated under vacuum. Purification by flash column
), 2.06 (s, 3H, OCOCH ), 2.04 (s, 3H, OCOCH ), 2.03 (s, 3H, chromatography (CH Cl /MeOH, 98:2 → 85:15, v/v) gave 38 (63 mg,
), 1.98 (s, 3H, OCOCH ), 1.91 (s, 3H, NHCOCH
); 13C NMR (126 48 μmol, 73%) as a white amorphous solid. To a solution of 38 (25
): δ 170.9, 170.7, 170.53, 170.48, 170.2, 170.14, 170.10, mg, 18.7 μmol) in dry MeOH (1 mL) was added methanolic sodium
3
(99 mg, 66 μmol) in water (660 μL). Then a solution of
N
2 3
2 2
S O
4
(108 mg, 0.53 mmol) in water (1.3 mL) was added and the
H-5’), 3.54 – 3.42 (m, 4H, H-2’, H-5, H-5’’, OCH
(
2
3
), 3.41 – 3.33 reaction was stirred for 3 hours. The mixture was then quenched
, extracted with AcOEt, dried over MgSO , filtered
2
2
S O
3
4
2
3
3
3
OCOCH
OCOCH
MHz, CDCl
3
3
3
3
2
2
3
3
3
1
1
1
69.3 (8 COCH
3
), 157.1 (ad, J = 37.3 Hz, NHCOCF
3
), 138.9, 138.3, methoxide (0.5 M) until pH = 8. The reaction was stirred at RT
+
38.2, 138.1 (4 CAr), 128.7, 128.53, 128.48, 128.31, 128.27, 128.1, overnight then quenched with Dowex 50WX8 H resin, filtered and
28.04, 127.99, 127.8, 127.7, 127.0 (20 CAr), 115.7 (ad, J = 287.7 Hz, concentrated in vacuo. The crude residue was purified by Sephadex
3
NHCOCF ), 102.4 (C-1’), 101.4 (C-1’’’), 100.2 (C-1’’), 99.6 (C-1), 80.4 P-2 size exclusion chromatography (Biogel® P-2, dH
2
O:n-BuOH, 99:1,
(
(
(
C-2’), 77.4 (C-4), 76.6 (C-3), 76.3 (C-3’), 75.7 (C-4’’), 75.1 (C-5), 75.0 v/v) to afford 1 (11.5 mg, 12.1 μmol, 70%) as a white amorphous
20
CH
2
Ph), 74.3 (CH
2
Ph), 73.7 (CH
2
Ph), 73.5 (CH
2
Ph), 72.9 (C-5’’), 72.5 solid. R
f
= 0.6, AcOEt/MeOH/AcOH/H
2
O 4:3:3:1; [훼]D = +10.1 (c 0.4,
1
C-5’), 72.0 (C-3’’), 71.0 (C-3’’’), 70.9 (C-5’’’), 69.8 (C-4’), 69.2 (C-2’’’),
H
2 2
O); H NMR (500 MHz, D O): δ 4.72 (m, 2H, H-1, H-1’’), 4.51 – 4.46
6
6
8.4 (OCH
2
CH
2
N
3
), 68.2 (C-6), 68.1 (C-6’), 66.8 (C-4’’’), 61.2 (C-6’’), (m, 2H, H-1’’’, H-1’), 4.17 (ad, J = 3.3 Hz, 1H, H-4’), 4.12 – 4.03 (m, 1H,
CH ), 23.7 OCHHCH ), 4.04 – 3.95 (m, 2H, H-6a, H-6b), 3.94 (ad, J = 3.4 Hz, 1H,
1.0 (C-6’’’), 57.0 (C-2), 54.6 (C-2’’), 50.7 (OCH
2
N
2 3
2 3
N
(
NHCOCH
3
), 20.9, 20.81, 20.79, 20.74, 20.72, 20.63, 20.55 (7 H-4’’’), 3.91 – 3.71 (m, 16H, H-2’’, H-2, H-6’a, H-6’b, H-6’’’a, H-6’’’b,
); 19F NMR (376 MHz, CDCl
) δ -75.97; HRMS (ESI ): m/z calcd H-6’’a, H-6’’b, H-3’’, H-3, H-5’’’, H-5’, H-4’’, H-4, H-3’, OCHHCH
+
N
),
OCOCH
for C72
3
3
2
3
+
H
86
F
3
5
N O
28: 1548.5309 [M+Na] ; found: 1548.5358.
3.68 (dd, J = 10.0, 3.4 Hz, 1H, H-3’’’), 3.66 – 3.52 (m, 4H, H-5’’, H-5, H-
’, H-2’’’), 3.52 – 3.44 (m, 2H, OCH CH ), 2.05 (s, 3H, NHCOCH );
O): δ 174.8 (NHCOCH ), 159.5 (ad, J = 37.6 Hz,
), 115.7 (ad, J = 286.3 Hz, NHCOCF ), 102.8, 102.9 (C-1’, C-
2
2
N
2 3
3
13
2
[
-Azidoethyl (2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl)-(14)-
3,6-di-O-acetyl-2-deoxy-2-(2,2,2-trichloroethoxycarbonylamino)-
C NMR (126 MHz, D
NHCOCF
2
3
3
3
β-D-glucopyranosyl]-(13)-(4-O-acetyl-2,6-di-O-benzyl-β-D-
1’’’), 102.7 (C-1’’), 100.2 (C-1), 82.0 (C-3’), 78.2, 78.1 (C-4, C-4’’), 75.3
(C-5’’’), 74.8 (C-5’, C-5’’), 74.5 (C-5), 72.4 (C-3’’’), 72.1, 71.7 (C-3, C-
galactopyranosyl)-(14)-3,6-di-O-benzyl-2-deoxy-2-
trifluoroacetamido-β-D-glucopyranoside (36) R
f
= 0.33, Tol/Acetone 3’), 70.9, 69.9 (C-2’, C-2’’’), 68.7 (OCH
); H NMR (500 MHz, CDCl ): δ 7.63 – 4’), 60.9, 60.9, 59.9, 59.8 (C-6, C-6’, C-6’’, C-6’’’), 55.51 (C-2’’), 55.12
.07 (m, 20H, HAr), 6.74 (d, J = 7.6 Hz, 1H, NHCOCF ), 5.38 – 5.32 (m, (C-2), 50.29 (OCH
H, H-4’, H-4’’’), 5.10 (dd, J = 10.5, 7.9 Hz, 1H, H-2’’’), 4.96 (dd, J = O): δ -75.8 (NHCOCF
0.5, 3.4 Hz, 1H, H-3’’’), 4.90 (d, J = 7.1 Hz, 1H, H-1), 4.86 – 4.73 (m, 894.2692 [M+Na] ; found: 894.2648.
H, CHHPh, CHHCCl , H-3’’), 4.71 – 4.38 (m, 12H, CHHCCl , CHHPh,
Ph, CHHPh, CHHPh, H-6’’a, H-1’’, H-1’’’, H-1’, 2-Azidoethyl
CCl ), 4.31 (d, J = 11.8 Hz, 1H, CHHPh), 4.14 – 3.95 (m, 6H, trifluoroacetamido-β-D-glucopyranosyl)-(13)-(β-D-
H-6’’’a, H-6’’’b, H-6’’b, OCHHCH , H-3, H-4), 3.86 (dd, J = 6.8, 1.2 galactopyranosyl)-(14)-2-acetamido-2-deoxy-β-D-
Hz, 1H, H-5’’’), 3.84 – 3.77 (m, 1H, H-6a), 3.76 – 3.34 (m, 12H, H-6b, glucopyranoside (2) To a solution of fully protected tetrasaccharide
H-3’, OCHHCH , H-2’’, H-2, H-5, H-5’, H-2’, H-5’’, OCH CHHN , H- 35 (100 mg, 65 μmol) in AcOEt (870 μL) was added a solution of
’a, H-6’b), 3.28 (ddd, J = 13.2, 5.4, 3.5 Hz, 1H, OCH CHHN ), 2.14 (s, NaBrO
H, OCOCH ), 2.08 (s, 3H, OCOCH ), 2.06 (s, 3H, OCOCH ), 2.05 (s, 3H, Na
), 2.02 (m, 6H, 2 OCOCH ), 1.96 (s, 3H, OCOCH
126 MHz, CDCl ): δ 170.6, 170.5, 170.4, 170.22, 170.17, 170.0, 169.3 with 10% Na
7 OCOCH ), 157.3 (q, J = 37.1 Hz, NHCOCF ), 154.2 (NHCO CH CCl ), and evaporated. Purification by flash column chromatography
38.3, 138.1, 138.0, 137.9 (4 CAr), 128.9, 128.6, 128.5, 128.4, 128.3, (CH Cl /MeOH, 98:2 → 85:15) gave 39 (54 mg, 46 μmol, 71%).
28.1, 128.0, 127.9, 127.9, 127.2 (20 CAr), 115.7 (ad, J = 288.4 Hz, Compound 39 (50 mg, 43 μmol) was dissolved in dry MeOH (2 mL)
2 2 3
CH N ), 68.47 (C-4’’’), 68.2 (C-
20
1
8
7
2
1
3
:2; [훼]D = +4.8 (c 1.0, CHCl
3
3
1
9
3
2
CH
2
N
3
), 22.11 (NHCOCH
3
); F NMR (282 MHz,
+
D
2
3
); HRMS (ESI ): m/z calcd for C30
H
F
48 3
5
N O
21
:
+
3
3
CHHPh, CH
NHCO CH
3
2
(β-D-galactopyranosyl)-(14)-(2-deoxy-2-
2
2
2 3
N
N
2 3
2
3
6
2
3
3
(99 mg, 0.66 mmol) in water (655 μL). Then a solution of
3
3
3
3
2 2
S O
4
(90 mg, 0.52 mmol) in water (1.3 mL) was added and the
13
OCOCH
3
3
3
); C NMR reaction was stirred for 6 hours. The mixture was then quenched
, extracted with AcOEt, dried over MgSO , filtered
(
(
3
2
2
S O
3
4
3
3
2
2
3
1
1
2
2
NHCOCF
CH CCl
5.4 (C-5’), 75.3 (CH
3.6 (CH Ph), 72.8 (C-5), 72.7 (C-5’’), 72.2 (C-3’’), 71.1 (C-3’’’), 70.9 chromatography (Biogel® P-2, dH
CH ), 68.2 (C-6, C-6’), compound 2 (20 mg, 23 μmol, 53%) as a white solid after freeze-dry.
6.8 (C-4’’’), 61.5 (C-6’’), 61.0 (C-6’’’), 56.4 (C-2’’), 55.9 (C-2), 50.7
3
), 102.8 (C-1’), 101.4 (C-1’’’), 101.2 (C-1’’), 99.2 (C-1), 95.7 and solid MeONa was added until pH = 9. The reaction was stirred
), 80.5 (C-2’), 76.6 (C-3’), 76.4, 76.3 (C-3, C-4), 76.2 (C-4’’), for 7 hours then quenched with Dowex 50WX8 H resin, filtered and
+
2
3
7
7
2
Ph), 74.4 (CH
2
Ph), 74.3 (CH
2 3
CCl ), 73.7 (CH
2
Ph), evaporated in vacuo. Crude product was purified by size exclusion
O:n-BuOH, 99:1, v/v) to obtain
2
2
(
6
(
C-5’’’), 69.7 (C-4’), 69.3 (C-2’’’), 68.5 (OCH
2
2 3
N
2
0
R
H
f
= 0.54, AcOEt/MeOH/AcOH/H
2
O 4:3:3:1; [훼]D = -10.7 (c 0.69,
1
OCH
2
CH
2
N
3
9
), 20.94, 20.88, 20.78, 20.75, 20.62, 20.58, 20.52 (7
2
O); H NMR (400 MHz, D
2
O) δ 4.85 (d, J = 7.7 Hz, 1H, H-1), 4.62 (d,
); 1 F NMR (376 MHz, CDCl
+
OCOCH
for C73
3
3
) δ -75.89; HRMS (ESI ): m/z calcd J = 8.1 Hz, 1H, H-1’’), 4.51 (d, J = 7.8 Hz, 1H, H-1’’’), 4.47 (d, J = 7.9 Hz,
+
H
85Cl
3
3
F N
5
O
29: 1680.4240 [M+Na] ; found: 1680.5447.
1H, H-1’), 4.20 (ad, J = 3.3 Hz, 1H, H-4’), 4.07 (ddd, J = 11.4, 5.5, 3.1
Hz, 1H, OCHHCH ), 4.01 (dd, J = 5.6, 2.3 Hz, 1H, H-6a), 3.98 (dd, J =
2 3
N
2
-Azidoethyl (β-D-galactopyranosyl)-(14)-(2-acetamido-2-deoxy- 5.7, 2.1 Hz, 1H, H-6’a), 3.95 (ad, J = 3.6 Hz, 1H, H-4’’’), 3.93 – 3.84 (m,
β-D-glucopyranosyl)-(13)-(β-D-galactopyranosyl)-(14)-(2-
4H, H-2, H-6b, H-6’b, H-3’’), 3.83 – 3.55 (m, 17H, H-2’’, H-6’’a, H-6’’b,
8
| J. Name., 2012, 00, 1-3
This journal is © The Royal Society of Chemistry 20xx
Please do not adjust margins

Page 9 of 15
O rP gl ea na is ce &d Bo i on mo to al e dc juu l sa tr mC haer mg i ins ts ry
Journal Name
H-6’’’a, H-6’’’b, OCHHCH
ARTICLE
2
N
3
, H-3’’’, H-5, H-5’’, H-5’, H-4, H-4’’, H-3’, 2-Azidoethyl
(2,6-di-O-benzyl-3,4-O-isopropylidene-β-D-
View Article Online
DOI: 10.1039/C8OB03066A
H-2’, H-2’’’, H-5’’’, H-3’), 3.55 – 3.40 (m, 2H, OCH
2
CH
O) δ 174.6 (NHCOCH
), 115.8 (ad, J = 286.5 Hz, NHCOCF
02.8 (C-1’, C-1’’’), 101.9 (C-1), 100.9 (C-1’’), 82.4 (C-3’), 78.4 (C-4’’), benzyl-2-deoxy-β-D-glucopyranoside (44) R
7.9 (C-4), 75.3, 74.8, 74.7, 74.6 (C-5, C-5’, C-5’’, C-5’’’), 72.5, 72.4, [훼]D = +5.3 (c 1.0, CHCl
1.5 (C-3’’, C-3’’’, C-3), 70.9 (C-2’’’), 69.8 (C-2’), 68.7 (OCH CH
2 3
N ), 2.06 (s, 3H, galactopyranosyl)-(14)-(3,6-di-O-benzyl-2-deoxy-2-
1
3
NHCOCH
3
); C NMR (126 MHz, D
2
3
), 159.5 (ad, trifluoroacetamido-β-D-glucopyranosyl)-(13)-(4-O-acetyl-2,6-di-
), 102.9, O-benzyl-β-D-galactopyranosyl)-(14)-2-acetamido-3,6-di-O-
= 0.65, Tol/Acetone 8:2;
); H NMR (500 MHz, CDCl ) δ 7.45 – 7.14 (m,
), 40H, HAr), 6.18 (d, J = 7.4 Hz, 1H, NHCOCF ), 5.79 (d, J = 7.4 Hz, 1H,
8.5 (C-4’’’), 68.1 (C-4’), 61.0, 60.9, 60.0, 59.7 (C-6, C-6’, C-6’’, C-6’’’), NHCOCH ), 5.42 (ad, J = 3.5 Hz, 1H, H-4’), 5.01 (d, J = 7.6 Hz, 1H, H-1),
); F NMR 4.96 – 4.89 (m, 2H, H-1’’, CHHPh), 4.82 (m, 2H, CHHPh, CHHPh), 4.78
(d, J = 11.0 Hz, 1H, CHHPh), 4.73 (d, J = 11.7 Hz, 1H, CHHPh), 4.63 (d,
J = 12.1 Hz, 1H, CHHPh), 4.61 – 4.54 (m, 3H, CH Ph, CHHPh), 4.61 –
.39 (m, 8H, H-1’’’, H-1’, 3 CH Ph), 4.33 (d, J = 11.8 Hz, 1H, CHHPh),
4.20 – 4.16 (m, 1H, H-4), 4.14 (dd, J = 5.6, 1.9 Hz, 1H, H-4’’’), 4.08 (at,
J = 6.1 Hz, 1H, H-3’’’), 4.06 – 3.96 (m, 3H, H-4’’, H-3, OCHHCH ),
0.019 M) together with 4Å molecular sieves. The mixture was stirred 3.88 (dd, J = 10.9, 4.3 Hz, 1H, H-6’’a), 3.78 – 3.63 (m, 9H, H-3’, H-3’’,
for 1 hour, then it was cooled to −30 °C and NIS (1.4 eq) was added H-5’’’, H-2’’, H-6’’b, OCHHCH , H-6’’’a, H-6’’’b, H-6’a), 3.57 (dd, J =
followed by TfOH (0.5 eq). The reaction was stirred for 90 minutes at 9.8, 6.3 Hz, 1H, H-6’b), 3.55 – 3.43 (m, 5H, OCH CHHN , H-5’, H-5, H-
the same temperature, then quenched with Et N, filtered over Celite 5’’, H-2’), 3.40 – 3.33 (m, 3H, H-2’’’, H-6a, H-6b), 3.30 – 3.21 (m, 2H,
and evaporated in vacuo. Crude was purified by flash column OCH CHHN , H-2), 1.99 (s, 3H, OCOCH ), 1.92 (s, 3H, NHCOCH ), 1.40
chromatography (see Supplementary Information). (s, 3H, CH ), 1.36 (s, 3H, (CH ) δ 170.7
); 13C NMR (126 MHz, CDCl
NHCOCH ), 170.1 (OCOCH ), 157.0 (ad, J = 36.9 Hz, NHCOCF ), 138.9,
(2,6-di-O-benzyl-3,4-O-isopropylidene-β-D- 138.6, 138.44, 138.42, 138.38, 138.21, 138.19, 138.1 (8 CAr), 128.6 –
), 110.0
), 102.5 (C-1’), 102.3 (C-1’’’), 99.7 (C-1’’), 99.6 (C-1), 80.6 (C-
2’’’), 80.5 (C-2’), 79.4 (C-3’’’), 77.6 (C-3’’), 77.3 (under CDCl peak, C-
J = 37.5 Hz, NHCOCF
3
3
1
7
7
6
5
f
20
1
3
3
2
N
2 3
3
3
19
2 2 3 3
5.7 (C-2), 54.9 (C-2’’), 50.3 (OCH CH N ), 22.2 (NHCOCH
+
(
8
376 MHz, D
94.2692 [M+Na] ; found: 894.2648.
2
O) δ -75.65; HRMS (ESI ): m/z calcd for C30
H
48
3
F N
5
O
21
:
+
2
4
2
General procedure for [2+2] glycosylation for compounds 43−45
Acceptor (1 eq) and donor (1.3−1.4 eq) were dissolved in dry CH
Cl
2 2
2 3
N
(
2 3
N
2
3
3
2
3
3
3
3
3
3
(
3
3
3
2
-Azidoethyl
galactopyranosyl)-(14)-(3,6-di-O-benzyl-2-deoxy-2-phthalimido- 127.6 (m, 39 CAr), 127.3 (CAr), 115.7 (ad, J = 288.7 Hz, NHCOCF
3
β-D-glucopyranosyl)-(13)-(4-O-acetyl-2,6-di-O-benzyl-β-D-
3 2
(C(CH )
galactopyranosyl)-(14)-2-acetamido-3,6-di-O-benzyl-2-deoxy-β-
3
2
0
D-glucopyranoside (43) R
f
= 0.86, Tol/Acetone 8:2; [훼] = +27.3 (c 4), 76.7 (C-3’), 76.6 (C-3), 76.3 (C-4’’), 75.5, 75.1, 75.0 (C-5’’, C-5,
D
1
1
.0, CHCl
3
); H NMR (500 MHz, CDCl
3
) δ 7.43 – 7.10 (m, 40H, HAr), 6.94 CH
2
Ph), 74.2 (CH
Ph, C-4’’’), 72.7 (C-5’), 72.2 (C-5’’’), 69.6 (C-4’), 69.1 (C-6’), 68.4,
CH ), 56.9 (C-2), 55.7 (C-2’’),
), 26.5 (CH
2
Ph), 73.8, 73.7, 73.51, 73.47, 73.4, 73.34, 73.30 (6
(
dd, J = 6.6, 2.9 Hz, 2H, HArNPhth), 6.90 – 6.84 (m, 2H, HArNPhth), 5.63 (d, CH
2
J = 7.4 Hz, 1H, NHCOCH
3
), 5.41 (ad, J = 3.6 Hz, 1H, H-4’), 5.29 (d, J = 68.3, 68.3, 68.1 (C-6’’, C-6, C-6’’’, OCH
CH ), 28.1 (CH
); 19F NMR (376 MHz, CDCl
2
2 3
N
8
.3 Hz, 1H, H-1’’), 4.88 (d, J = 7.6 Hz, 1H, H-1), 4.85 – 4.79 (m, 3H, 50.7 (OCH
2
2
N
3
3
3
), 23.7 (NHCOCH
3
), 20.8
+
CHHPh, CHHPh, CHHPh), 4.75 (d, J = 11.7 Hz, 1H, CHHPh), 4.65 (d, J = (OCOCH
3
3
) δ -75.72; HRMS (ESI ): m/z calcd
+
1
7
4
2.0 Hz, 1H, CHHPh), 4.53 (d, J = 12.1 Hz, 1H, CHHPh), 4.49 – 4.37 (m, for C91
H, H-1’’’, CHHPh, CHHPh, CHHPh, CHHPh, CHHPh, CHHPh), 4.36 –
.26 (m, 3H, H-3’’, CHHPh, CHHPh), 4.26 – 4.13 (m, 3H, H-1’, CHHPh, 2-Azidoethyl
H
102
F
3
N
5
O
22: 1696.6861 [M+Na] ; found: 1696.5470.
(2,6-di-O-benzyl-3,4-O-isopropylidene-β-D-
H-2’’), 4.13 – 4.01 (m, 5H, H-4’’, CHHPh, H-4’’’, H-3’’’, H-3), 3.96 – 3.89 galactopyranosyl)-(14)-[3,6-di-O-benzyl-2-deoxy-2-(2,2,2-
(
(
6
3
5
OCH
3
(
1
C
1
1
1
7
7
(
(
6
m, 2H, H-6a, OCHHCH
2 3
N ), 3.84 (at, J = 8.5 Hz, 1H, H-4), 3.82 – 3.78 trichloroethoxycarbonylamino)-β-D-glucopyranosyl]-(13)-(4-O-
m, 1H, H-6b), 3.75 (dd, J = 6.4, 2.0 Hz, 1H, H-5’’’), 3.65 (dd, J = 9.9, acetyl-2,6-di-O-benzyl-β-D-galactopyranosyl)-(14)-3,6-di-O-
.5 Hz, 1H, H-6’’’a), 3.62 – 3.54 (m, 3H, H-5’’, H-6’’’b, OCHHCH
.54 – 3.47 (m, 2H, H-3’, H-6a’’), 3.47 – 3.28 (m, 7H, OCH CHHN
’, H-2’’’, H-2’, H-6’’b, H-6’a, H-6’b), 3.24 – 3.11 (m, 3H, H-5, H-2, CDCl
CHHN ), 2.01 (s, 3H, OCOCH ), 1.85 (s, 3H, NHCOCH ), 1.37 (s, 5.43 (ad, J = 3.6 Hz, 1H, H-4’), 4.90 (d, J = 7.0 Hz, 1H, H-1), 4.87 – 4.77
H, CH ), 1.33 (s, 3H, CH ) δ 170.6 (m, 4H, CHHPh, CHHPh, CHHPh, NHCO CH CCl ), 4.74 – 4.28 (m, 20H,
); 13C NMR (126 MHz, CDCl
NHCOCH ), 170.1 (OCOCH ), 167.6 (2 CONPhth), 139.0, 138.9, 138.8, H-1’’, H-1’’’, H-1’, CHHCCl , 8 CHHPh), 4.13 (dd, J = 5.5, 2.0 Hz, 1H, H-
38.6, 138.5, 138.4, 138.3, 138.2 (8 CAr), 133.6 (2 CNPhth), 131.4 (2 4’’’), 4.10 – 3.93 (m, 5H, H-3, H-3’’’, H-4, H-4’’, OCHHCH ), 3.82
NPhth), 128.50, 128.46, 128.44, 128.39, 128.2, 128.11, 128.08, (atd, J = 10.4, 9.8, 4.3 Hz, 1H, H-6’’a), 3.79 – 3.76 (m, 1H, H-6’’b), 3.73
28.03, 127.92, 127.87, 127.8, 127.69, 127.67, 127.60, 127.58, 127.5, (ddd, J = 12.9, 5.3, 2.0 Hz, 1H, H-5’’’), 3.70 – 3.60 (m, 5H, OCHHCH
27.0, 127.0, 126.6 (40 HAr) 123.1 (2 CNPhth), 109.8 (C(CH ), 102.5 (C- H-6a, H-6b, H-6’’’a, H-3’), 3.59 – 3.49 (m, 5H, H-2, H-6’’’b, H-5’, H-5,
’’’), 102.3 (C-1’), 99.5 (C-1), 99.2 (C-1’’), 80.7 (C-2’’’), 79.5 (C-3’), H-2’), 3.49 – 3.34 (m, 7H, OCH CHHN , H-2’’, H-6’a, H-6’b, H-5’’, H-3’’,
9.3, 78.8, 78.0, 77.4 (C-3’’’, C-4’’’, C-3, C-2’), 77.0 (C-3’’), 76.4 (C-4), H-2’’’), 3.28 (ddd, J = 13.2, 5.4, 3.5 Hz, 1H, OCH CHHN ), 2.02 (s, 3H,
5.4 (C-5’’), 74.9 (C-5), 74.5 (CH Ph), 74.4 (CH Ph), 74.2 (CH Ph), 74.0 OCOCH ), 1.41 (s, 3H, CH ), 1.36 (s, 3H, CH
); 13C NMR (101 MHz,
Ph), 73.65 (CH Ph), 73.5 (C-4’’), 73.3 (CH Ph), 73.2 CDCl ) δ 170.0 (OCOCH ), 157.1 (ad, J = 37.2 Hz, NHCOCF ), 153.9
Ph), 72.9 (C-5’), 72.1 (C-5’’’), 70.2 (C-4’), 69.2, 68.9, 68.3, 68.0, (NHCO CH CCl ), 138.8, 138.7, 138.5, 138.12, 138.06, 138.0 (8 CAr),
CH , C-6, C-6’, C-6’’, C-6’’’), 57.0 (C-2), 56.2 (C-2’’), 50.7 128.7, 128.6, 128.53, 128.47, 128.43, 128.41, 128.37, 128.33, 128.30,
2 3 f
N ), benzyl-2-deoxy-2-trifluoroacetamido-β-D-glucopyranoside (45) R =
2
0
1
2
3
, H- 0.46, Tol/Acetone 9:1; [훼]D = +5.6 (c 1.0, CHCl
3
); H NMR (500 MHz,
),
3
) δ 7.39 – 7.21 (m, 40H, HAr), 6.72 (d, J = 7.6 Hz, 1H, NHCOCF
3
2
3
3
3
3
3
3
2
2
3
3
3
3
2 3
N
2 3
N ,
3 2
)
2
3
2
3
2
2
2
3
3
3
CH
CH
2
Ph), 73.68 (CH
2
2
2
3
3
3
2
2
2
3
7.9 (OCH
2
2 3
N
(
OCH
2
CH
2
N
3
), 28.1 (CH
3
), 26.5 (CH
3
), 23.7 (NHCOCH
3
), 20.9 (OCOCH
3
); 128.2, 128.1, 128.04, 127.99, 127.9, 127.8, 127.68, 127.67, 127.63,
), 109.9
), 102.9 (C-1’), 102.2 (C-1’’’), 101.2 (C-1’’), 99.2 (C-1), 95.8
+
+
HRMS (ESI ): m/z calcd for C97
1
H
105
5
N O
23; 1730.7098 [M+Na] ; found: 127.56, 127.2 (40 CAr), 115.6 (ad, J = 288.6 Hz, NHCOCF
3
730.7119.
(C(CH
3 2
)
(
CH CCl
2
3
), 80.7 (C-2’’’), 80.4 (C-2’), 79.5 (C-3’’’), 79.2 (C-3’’), 77.4 (C-
This journal is © The Royal Society of Chemistry 20xx
J. Name., 2013, 00, 1-3 | 9
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O rP gl ea na is ce &d Bo i on mo to al e dc juu l sa tr mC haer mg i ins ts ry
Page 10 of 15
ARTICLE
Journal Name
3
7
7
6
2
’), 76.4, 76.3 (C-3, C-4, C-4’’), 75.5, 75.4 (C-5, C-5’’), 75.0, 74.33,
4.25, 73.8, 73.5, 73.4, 73.3 (8 CH Ph, CH CCl , C-4’’’), 73.1 (C-5’), 2-Azidoethyl
2.1 (C-5’’’), 69.8 (C-4’), 69.1 (C-6’’’), 68.5 (OCH CH
CH
View Article Online
DOI: 10.1039/C8OB03066A
(4-O-acetyl-2,6-di-O-benzyl-β-D-galactopyranosyl)-
2
2
3
2
2
N
3
), 68.3, 68.2 (C- (14)-[3,6-di-O-benzyl-2-deoxy-2-(2,2,2-
), 28.1 (CH
, C-6’, C-6’’), 57.0 (C-2’’), 55.8 (C-2), 50.7 (OCH
2
2
N
3
3
), trichloroethoxycarbonylamino)-β-D-glucopyranosyl]-(13)-(4-O-
); 9F NMR (376 MHz, CDCl
1
) δ -75.88; HRMS acetyl-2,6-di-O-benzyl-β-D-galactopyranosyl)-(14)-3,6-di-O-
6.5 (CH
+
3
), 20.8 (OCOCH
3
3
+
(
ESI ): m/z calcd for C92
H
101Cl
F
3 3
5
N O
23: 1828.5797 [M+Na] ; found benzyl-2-deoxy-2-trifluoroacetamido-β-D-glucopyranoside (51) R
f
=
20
1
1
828.4799.
0.75. Tol/Acetone 7:3; [훼]D = +1.9 (c 1.0, CHCl
CDCl ) δ 7.38 – 7.17 (m, 40H, HAr), 6.74 (d, J = 7.6 Hz, 1H, NHCOCF
General procedure for the formation of acceptors 50−52 and 55 via 5.45 (ad, J = 3.6 Hz, 1H, H-4’), 5.35 (ad, J = 3.5 Hz, 1H, H-4’’’), 4.90 (d,
’,4’-O-isopropylidene cleavage, orthoester formation, and J = 7.1 Hz, 1H, H-1), 4.89 – 4.80 (m, 3H, CHHPh, CHHPh, CHHCCl ),
rearrangement sequence: 4.78 (d, J = 7.9 Hz, 1H, H-1’’), 4.73 – 4.38 (m, 16H, 6 CH Ph, CHHCCl
Fully protected tetrasaccharide (1 eq) was stirred in aq. 80% AcOH H-1’’’, H-1’, NHCO CH CCl ), 4.34 (d, J = 11.7 Hz, 1H, CHHPh), 4.24 (d,
0.02 M) for 4-5 hours at 60−70 °C. After dilution with toluene the J = 11.9 Hz, 1H, CHHPh), 4.08 (at, J = 8.1 Hz, 1H, H-4), 4.04 – 3.96 (m,
solvents were removed and crude was purified by column 3H, H-3, H-4’’, OCHHCH ), 3.82 (dd, J = 11.0, 4.4 Hz, 1H, H-6’’a),
chromatography to afford the corresponding diol derivative. Diol (1 3.80 – 3.74 (m, 2H, H-6’’b, H-6a), 3.71 – 3.60 (m, 4H, H-6b,
eq) was then dissolved in CH CN (0.05 M) and reacted with OCHHCH , H-3’’’, H-3’), 3.59 – 3.51 (m, 6H, H-2, H-5’’’, H-5’, H-5, H-
CH C(OCH (3 eq) and p-TsOH (0.1-0.5 eq). After 30 minutes the 3’’, H-2’), 3.47 – 3.40 (m, 5H, OCH CHHN , H-2’’, H-6’a, H-5’’, H-2’’’),
reaction was quenched with Et N and concentrated to dryness. 3.40 – 3.34 (m, 1H, H-6’b), 3.33 – 3.25 (m, 3H, OCH CHHN , H-6’’’a,
Crude orthoester product was then dissolved in aq. 80% AcOH (0.05 H-6’’’b), 2.03 – 2.00 (m, 6H, 2 OCOCH ); C NMR (126 MHz, CDCl ) δ
M) and stirred for 1 hour at RT. After solvent removal, crude was 170.90 (OCOCH ), 169.87 (OCOCH ), 157.1 (ad, J = 37.3 Hz,
purified by flash column chromatography (see Supplementary NHCOCF ), 153.8 (NHCO CH Cl ), 138.61, 138.60, 138.23, 138.16,
Information). 138.0, 137.91, 137.88, 137.8 (8 CAr), 129.0, 128.6, 128.54, 128.48,
28.43, 128.39, 128.38, 128.36, 128.3, 128.0, 127.92, 127.89, 127.87,
(4-O-acetyl-2,6-di-O-benzyl-β-D-galactopyranosyl)- 127.84, 127.81, 127.75, 127.71, 127.69, 127.65, 127.57, 127.4 (40
Ar), 115.6 (ad, J = 288.2 Hz, NHCOCF ), 102.7 (C-1’), 102.5 (C-1’’’),
100.9 (C-1’’), 99.0 (C-1), 95.6 (CH CCl ), 80.2 (C-2’), 80.1 (C-2’’’), 79.2
(C-3’’), 77.7 (C-3’), 76.3, 76.2 (C-3, C-4, C-4’’), 75.3, 75.2 (C-5, C-5’’),
Ph, CH CCl ), 72.9 (C-5’),
CH ),
), 5.42 (ad, 68.1, 68.0 (C-6, C-6’, C-6’’), 67.3 (C-6’’’), 57.0 (C-2’’), 55.7 (C-2), 50.6
3
); H NMR (500 MHz,
3
3
),
3
3
2
3
,
2
2
3
(
2 3
N
3
2 3
N
3
3
)
3
2
3
3
2
3
13
3
3
3
3
3
2
2
3
1
2
-Azidoethyl
(
14)-(3,6-di-O-benzyl-2-deoxy-2-trifluoroacetamido-β-D-
C
3
glucopyranosyl)-(13)-(4-O-acetyl-2,6-di-O-benzyl-β-D-
2
3
galactopyranosyl)-(14)-2-acetamido-3,6-di-O-benzyl-2-deoxy-β-
2
0
D-glucopyranoside (50) R
f
= 0.53, Tol/Acetone 8:2; [훼] = -2.8 (c 1.0, 75.11, 74.12, 73.6, 73.4, 73.3, 73.2 (8 CH
2
2
3
D
1
CHCl
3
); H NMR (500 MHz, CDCl
3
) δ 7.38 – 7.20 (m, 40H, HAr), 6.26 (d, 72.4 (C-3’’’), 72.0 (C-5’’’), 69.6 (C-4’), 69.6 (C-4’’’), 68.3 (OCH
), 5.76 (d, J = 7.4 Hz, 1H, NHCOCH
J = 3.6 Hz, 1H, H-4’), 5.35 (dd, J = 3.5, 1.1 Hz, 1H, H-4’’’), 5.00 (d, J = (OCH
.8 Hz, 1H, H-1), 4.97 (d, J = 7.1 Hz, 1H, H-1’), 4.91 (d, J = 11.1 Hz, 1H, CDCl
2
2 3
N
J = 8.2 Hz, 1H, NHCOCF
3
3
1
9
2
CH
2
N
3
), 20.74 (OCOCH
-75.89 (NHCOCF
3
), 20.68 (OCOCH
); HRMS (ESI ): m/z calcd for
3
); F NMR (376 MHz,
+
7
3
)
δ
3
+
CHHPh), 4.85 (d, J = 11.3 Hz, 1H, CHHPh), 4.88 – 4.71 (m, 2H, CHHPh,
CHHPh), 4.69 (d, J = 11.3 Hz, 1H, CHHPh), 4.64 (d, J = 12.1 Hz, 1H,
91 3 3 5
C H99Cl F N O24: 1830.5595 [M+Na] ; found: 1830.5514.
CHHPh), 4.60 – 4.26 (m, 12H, H-1’’’, H-1’, CH
CH Ph, CH
H, H-4’’), 4.03 – 3.96 (m, 2H, OCHHCH
.3 Hz, 1H, H-6’’a), 3.79 – 3.62 (m, 8H, H-6’’b, H-6a, H-6b, H-3’’’, H-3’, galactopyranosyl)-(14)-3,6-di-O-benzyl-2-deoxy-2-
), 3.60 – 3.55 (m, 1H, H-5’’’), 3.55 – 3.43 (m, trifluoroacetamido-β-D-glucopyranoside (52) R = 0.43, Tol/Acetone
); 1H NMR (500 MHz, CDCl
) δ 7.39 –
.37 (m, 2H, H-6’a, H-6’b), 3.33 (m, 2H, H-6’’’a, H-6’’’b), 3.25 3.28 – 7.21 (m, 40H, HAr), 6.82 (d, J = 7.6 Hz, 1H, NHCOCF ), 5.41 (ad, J = 3.5
.21 (m, 2H, OCH CHHN , H-2), 2.02 (s, 3H, OCOCH ), 1.99 (s, 3H, Hz, 1H, H-4’), 5.35 (ad, J = 3.5 Hz, 1H, H-4’’’), 5.26 (d, J = 7.8 Hz, 1H,
), 1.91 (s, 3H, NHCOCH ) δ 170.9, NHCOCH ), 5.06 (d, J = 7.5 Hz, 1H, H-1’’), 4.91 – 4.84 (m, 3H, H-1,
); 13C NMR (126 MHz, CDCl
70.6, 169.9 (3 COCH ), 156.9 (ad, J = 36.8 Hz, NHCOCF ), 138.8, CH Ph), 4.83 – 4.76 (m, 2H, CH Ph), 4.70 – 4.60 (m, 3H, CH Ph,
38.4, 138.12, 138.08, 138.07, 138.06, 137.9, 137.7 (8 CAr), 129.02, CHHPh), 4.59 – 4.52 (m, 3H, CHHPh, CH Ph), 4.51 (d, J = 7.8 Hz, 1H,
28.96, 128.59, 128.58, 128.56, 128.41, 128.38, 128.37, 128.35, H-1’’’), 4.48 – 4.38 (m, 5H, H-1’, 2 CH Ph), 4.34 (d, J = 11.7 Hz, 1H,
28.22, 128.17, 128.14, 128.09, 128.0, 127.87, 127.85, 127.81, CHHPh), 4.28 (d, J = 12.1 Hz, 1H, CHHPh), 4.11 – 3.94 (m, 5H, H-3’’, H-
27.75, 127.7, 127.64, 127.55, 127.2 (40 CAr), 115.6 (ad, J = 288.6 Hz, 3, H-4, H-4’’, OCHHCH ), 3.86 – 3.75 (m, 3H, H-6’’a, H-6’’b, H-6a),
), 102.8 (C-1’’’), 102.4 (C-1’), 99.5 (C-1), 99.4 (C-1’’), 80.3 (C- 3.71 (dd, J = 10.6, 3.5 Hz, 1H, H-6b), 3.68 – 3.50 (m, 9H, OCHHCH
’), 79.9 (C-2’’’), 77.3 (C-3), 77.2 (C-3’’), 76.7 (C-3’), 76.4 (C-4), 76.2 H-2, H-6b, H-3’’’, H-3’, H-5’’’, H-5’, H-5’’, H-2’), 3.47 – 3.39 (m, 3H,
C-4’’), 75.4 (C-5’’), 75.1 (CH Ph), 74.93, 74.86 (CH Ph, C-5’), 74.1, OCH CHHN , H-6’a, H-2’’’), 3.39 – 3.33 (m, 4H, H-2’’, H-6’b, H-6’’’a, H-
3.6, 73.46, 73.45, 73.24, 73.19 (6 CH Ph), 72.6 (C-5’), 72.3 (C-3’’’), 6’’’b), 3.29 (m, 1H, OCH CHHN ), 2.01 (s, 6H, 2 OCOCH ), 1.52 (s, 3H,
2.1 (C-5’’’), 69.54 (C-4’’’), 69.47 (C-4’), 68.3 (OCH ); C NMR (126 MHz, CDCl ) δ 170.9, 170.2, 170.0 (3
), 157.1 (ad, J = 37.2 Hz, NHCOCF ), 138.9, 138.7, 138.3, 138.1,
137.99, 137.95, 137.9, 137.8 (8 CAr), 128.6, 128.43, 128.41, 128.36,
2
Ph, CH
Ph), 4.18 (dd, J = 9.5, 8.2 Hz, 1H, H-3), 4.05 (at, J = 8.2 Hz, (14)-(2-acetamido-3,6-di-O-benzyl-2-deoxy-β-D-
, H-4), 3.84 (dd, J = 10.9, glucopyranosyl)-(13)-(4-O-acetyl-2,6-di-O-benzyl-β-D-
2
Ph, CH
2
Ph, 2-Azidoethyl
(4-O-acetyl-2,6-di-O-benzyl-β-D-galactopyranosyl)-
2
2
1
4
2 3
N
H-3’’, H-2’’, OCHHCH
2
N
3
f
20
5
3
3
H, H-5’, H-5, H-5’’, H-2’, OCH
2
CHHN
3
), 3.43 – 3.40 (m, 1H, H-2’’’), 7:3; [훼]D = +3.6 (c 1.0, CHCl
3
3
3
2
3
3
OCOCH
3
3
3
3
1
1
1
1
1
3
3
2
2
2
2
2
2 3
N
NHCOCF
3
2 3
N ,
2
(
2
2
2
3
7
7
2
2
3
3
13
2
CH
2 3
N ), 68.1, 68.0 NHCOCH
3
3
(
(
C-6, C-6’, C-6’’), 67.3 (C-6’’’), 56.8 (C-2), 55.5 (C-2’’), 50.6 COCH
3
3
19
OCH
2
CH
2 3
N
), 23.6 (NHCOCH
3
), 20.7 (OCOCH
3
), 20.6 (OCOCH
3
);
F
+
NMR (376 MHz, CDCl
for C90
3
) δ -75.72 (NHCOCF
3
); HRMS (ESI ): m/z calcd 128.24, 128.22, 128.20, 128.12, 128.08, 127.9, 127.8, 127.71, 127.69,
+
H
100
F
3
N
5
O
23; 1698.6659 [M+Na] ; found 1698.6711.
127.63, 127.55, 127.5, 127.2 (40 CAr), 115.6 (ad, J = 288.6 Hz,
1
0 | J. Name., 2012, 00, 1-3
This journal is © The Royal Society of Chemistry 20xx
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Page 11 of 15
O rP gl ea na is ce &d Bo i on mo to al e dc juu l sa tr mC haer mg i ins ts ry
Journal Name
), 102.8 (C-1’), 102.5 (C-1’’’), 100.2 (C-1’’), 99.0 (C-1), 80.0 1:1; [훼]D = +3.8 (c 1.0, CHCl
ARTICLE
20
1
NHCOCF
C-2’’’), 79.5 (C-2’), 78.8 (C-3’), 78.0 (C-3’’), 76.4, 76.3, 76.1 (C-3, C-4, (m, 40H, HAr), 6.32 (d, J = 8.3 Hz, 1H, NHCOCF
C-4’’), 75.3 (C-5), 75.09 (C-5’’), 75.06 (CH Ph), 75.0 (CH Ph), 73.9 1H, NHCOCH ), 5.39 (ad, J = 3.6 Hz, 1H, H-4’), 5.37 (br, 1H, H-4’’’),
Ph), 73.3 (CH Ph), 73.2 (CH Ph), 5.36 – 5.34 (m, 1H, H-4’’’’’), 5.10 (dd, J = 10.4, 7.9 Hz, 1H, H-2’’’’’),
3.0 (C-5’), 72.3 (C-3’’’), 72.0 (C-5’’’), 69.54 (C-4’’’), 69.46 (C-4’), 68.6 5.01 – 4.88 (m, 4H, H-3’’’’’, CHHPh, H-1, H-1’’), 4.86 – 4.73 (m, 5H, H-
CH , C-6), 68.2 (C-6’’), 67.4 (C-6’’’), 56.6 (C-2’’), 3’’’’, CHHPh, CH Ph, NHCO CH CCl ), 4.68 (m, 2H, CHHCCl , CHHPh),
CH ), 23.2 (NHCOCH ), 20.8 (2 OCOCH ); 4.63 – 4.53 (m, 7H, H-6’’’’a, CHHCCl , CH Ph, CH Ph , H-1’’’’, CHHPh),
23; 1698.6654 [M+Na] ; 4.52 – 4.39 (m, 7H, H-1’’’’’, H-1’’’, CHHPh, 2 CH Ph), 4.41 – 4.33 (m,
2H, CHHPh, H-1’), 4.33 – 4.27 (m, 2H, CH Ph), 4.16 (dd, J = 9.6, 8.2 Hz,
H, H-3), 4.12 – 4.09 (m, 2H, H-6’’’’’a, H-6’’’’’b), 4.08 – 3.96 (m, 4H,
(4-O-acetyl-2,6-di-O-benzyl-β-D-galactopyranosyl)- H-6’’’’b, OCHHCH , H-4’’, H-4), 3.88 (dd, J = 6.8, 1.3 Hz, 2H, H-5’’’’’),
3.82 – 3.56 (m, 12H, H-2’’, H-2’’’’, H-6’’a, H-6’’b, H-6a, H-6b,
OCHHCH , H-5’’’’, H-3’’, H-3’, H-3’’’, H-4’’’’), 3.54 – 3.42 (m, 6H, H-
5’’’, H-5’, H-5 H-2’, H-2’’’, OCH CHHN ), 3.41 – 3.31 (m, 5H, H-6’’’a,
3
3
); H NMR (500 MHz, CDCl
3
) δ 7.35 – 7.20
View Article Online
DOI: 10.1039/C8OB03066A
(
3
), 5.80 (d, J = 7.4 Hz,
2
2
3
(
CH
2
Ph), 73.6 (2 CH
2
Ph), 73.4 (CH
2
2
2
7
(
C-6’), 68.3 (OCH
2
N
2 3
2
2
2
3
3
5
5.4 (C-2), 50.6 (OCH
2
2
N
3
3
3
+
3
2
2
+
HRMS (ESI ): m/z calcd for C90
found 1698.4514.
H
100
F
3 5
N O
2
2
1
2
-Azidoethyl
2 3
N
(
14)-(2-acetamido-3,6-di-O-benzyl-2-deoxy-β-D-
glucopyranosyl)-(13)-(4-O-acetyl-2,6-di-O-benzyl-β-D-
2 3
N
galactopyranosyl)-(14)-2-acetamido-3,6-di-O-benzyl-2-deoxy-β-
,
2
3
2
0
D-glucopyranoside (55) R
f
= 0.23, cHex/Acetone 7:3; [훼] = +4.9 (c H-6’’’b, H-6’a, H-6’b, H-5’’), 3.27 – 3.20 (m, 2H, H-2, OCH
2
CHHN
), 2.08 (s, 3H, OCOCH
), 2.02 (s, 3H, OCOCH
3
),
3
),
3
),
D
1
1
.0, CHCl
3
); H NMR (500 MHz, CDCl
3
) δ 7.38 – 7.18 (m, 40H, HAr), 5.74 2.15 (s, 3H, OCOCH
), 5.41 (ad, J = 3.5 Hz, 1H, H-4’), 5.34 (dd, 2.06 (s, 3H, OCOCH
J = 3.5, 1.0 Hz, 1H, H-4’’’), 5.22 (d, J = 7.9 Hz, 1H, NHCOCH
.98 (m, 2H, H-1, H-1’’), 4.91 (d, J = 11.1 Hz, 1H, CHHPh), 4.88 – 4.84 MHz, CDCl
m, 2H, CHHPh, CHHPh), 4.81 (d, J = 11.6 Hz, 1H, CHHPh), 4.68 – 4.60 169.9, 169.2 (9 COCH
m, 3H, 2 CH Ph, CHHPh), 4.59 – 4.52 (m, 2H, CH Ph), 4.50 (d, J = 7.8 (NHCO CH CCl ), 138.8, 138.4, 138.3, 138.12, 138.07, 138.0, 137.9,
Ph), 4.31 (d, J = 11.9 Hz, 137.8 (8 CAr), 128.7, 128.4, 128.4 – 128.2, 128.19, 128.18, 128.15,
H, CHHPh), 4.26 (d, J = 12.0 Hz, 1H, CHHPh), 4.19 (dd, J = 9.5, 8.1 Hz, 128.1, 128.0, 127.87, 127.85, 127.8, 127.71, 127.65, 127.6, 127.5,
H, H-3), 4.06 – 3.95 (m, 4H, H-3’’, H-4, H-4’’, OCHHCH ), 3.83 (dd, 127.2, 127.1 (40 CAr), 115.6 (ad, J = 288.7 Hz, NHCOCF ), 102.4 (C-
3
), 2.09 (s, 3H, OCOCH
3
(
d, J = 7.4 Hz, 1H, NHCOCH
3
3
), 2.03 (s, 3H, OCOCH
3
1
3
3
’’), 5.03 – 1.99 – 1.96 (m, 6H, 2 OCOCH
3
3
), 1.91 (s, 3H, NHCOCH ); C NMR (101
4
(
(
3
) δ 170.7, 170.54, 170.45, 170.4, 170.2, 170.1, 170.0,
), 156.9 (ad, J = 36.9 Hz, NHCOCF ), 154.1
3
3
2
2
2
2
3
Hz, 1H, H-1’’’), 4.47 – 4.36 (m, 5H, H-1’, 2 CH
2
1
1
2
N
3
3
J = 10.8, 4.0 Hz, 1H, H-6a), 3.79 – 3.73 (m, 2H, H-6b, H-6’’a), 3.69 – 1’’’), 102.3 (C-1’), 101.2 (C-1’’’’’), 101.0 (C-1’’’’), 99.5 (C-1), 99.3 (C-
3
5
.60 (m, 4H, H-6’’b, OCHHCH
’’’, H-5’, H-5’’, H-2’), 3.48 – 3.30 (m, 7H, OCH
2
N
3
, H-3’’’, H-3’), 3.58 – 3.48 (m, 4H, H- 1’’), 95.6 (CH
CHHN , H-2’’, H-6’’’a, C-3’), 76.4 (C-4), 76.1 (C-3’’), 75.8 (C-4’’), 75.3 (C-5’’), 75.1, 74.9, 74.8,
CHHN ), 74.3, 74.1, 73.7, 73.51, 73.48, 73.3, 73.2 (8 CH Ph, C-5, CH CCl ), 72.5
), 1.49 (s, 3H, (C-5’, C-5’’’, C-5’’’’), 72.3 (C-3’’’’), 71.0 (C-3’’’’’), 70.7 (C-5’’’’’), 69.7,
) δ 170.8 (OCOCH ), 170.6 69.5 (C-4’, C-4’’’), 69.2 (C-2’’’’’), 68.2, 68.1, 68.0, 67.8 (C-6’’, C-6, C-6’,
), 169.9 (OCOCH ), 138.9, 138.8, 138.4, C-6’’’, OCH CH ), 66.66 (C-4’’’’’), 61.4 (C-6’’’’), 60.9 (C-6’’’’’), 56.8
38.2, 138.1, 137.8 (8 CAr), 128.6 – 127.0 (m, 40 CAr), 102.5 (C-1’’’), (C-2), 56.3 (C-2’’’’), 55.8 (C-2’’), 50.6 (OCH CH ), 23.5 (NHCOCH ),
02.4 (C-1’), 100.3 (C-1’’), 99.5 (C-1), 80.0 (C-2’’’), 79.7 (C-2’), 78.8 (C- 20.79, 20.77, 20.7, 20.63, 20.61, 20.5 (8 OCOCH
2 3
CCl ), 80.3 (C-2’, C-2’’’), 77.3 (C-3), 76.7 (C-4’’’’, C-3’’’,
2
3
H-6’’’b, H-6’a, H-6’b, H-2’’’), 3.28 – 3.21 (m, 2H, H-2, OCH
.01 – 1.97 (m, 6H, 2 OCOCH ), 1.89 (s, 3H, NHCOCH
NHCOCH
); 13C NMR (126 MHz, CDCl
), 170.2 (NHCOCH
2
3
2
2
3
2
3
3
3
3
3
(
NHCOCH
3
3
3
2
2 3
N
1
1
3
7
7
2
2
N
3
3
); 19F NMR (376 MHz,
3
’), 78.1 (C-4’’), 77.4 (under CDCl
5.11, 75.06, 75.0, 74.9 (2 CH Ph, C-5’’, C-5), 74.1 (CH
3.3, 73.2 (5 CH Ph), 72.8 (C-5’), 72.3 (C-3’’’), 72.0 (C-5’’’), 69.6 (C-4’,
CH
3
peak, C-3’’), 76.6 (C-3), 76.2 (C-4), CDCl
3
)
δ
-75.69 (NHCOCF
3
); HRMS (ESI+): m/z calcd for
+
2
2
Ph), 73.6, 73.4,
C
117
H
134Cl
3
F
3
6
N O
40: 2447.7546 [M+Na] ; found: 2447.8903.
2
C-4’’’), 68.6 (C-6’), 68.3, 68.2 (C-6, C-6’’, OCH
5
2
2
N
3
), 67.4 (C-6’’’), 2-Azidoethyl (2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl)-(14)-
6.7 (C-2, C-2’’), 50.6 (OCH
2
CH
N
2 3
), 23.6 (NHCOCH
3
), 23.1 (3,6-di-O-acetyl-2-deoxy-2-trifluoroacetamido-β-D-
+
(
NHCOCH
3
), 20.8 (OCOCH
3
), 20.7 (OCOCH
3
); HRMS (ESI ): m/z calcd glucopyranosyl)-(13)-(4-O-acetyl-2,6-di-O-benzyl-β-D-
+
for C90
H
103
N
5
O
23: 1644.6942 [M+Na] ; found: 1644.7008.
galactopyranosyl)-(14)-(2-acetamido-3,6-di-O-benzyl-2-deoxy-
β-D-glucopyranosyl)-(13)-(4-O-acetyl-2,6-di-O-benzyl-β-D-
General procedure for [2+4] glycosylation for compounds 56, 57, 59 galactopyranosyl)-(14)-2-acetamido-3,6-di-O-benzyl-2-deoxy-β-
Acceptor (1 eq) and donor (1.5 eq) were dissolved in dry CH
2
0
2
Cl
2
D-glucopyranoside (57) R
f
= 0.40, cHex/Acetone 1:1; [훼] = +5.4 (c
D
1
(
0.015 M) together with 4Å molecular sieves. The mixture was stirred 1.0, CHCl
3
); H NMR (500 MHz, CDCl
3
) δ 7.46 – 7.10 (m, 40H, HAr), 6.18
), 5.72 (d, J = 7.4 Hz, 1H, NHCOCH ), 5.40
was added followed by TfOH (0.8 eq). The reaction was stirred for 1 – 5.34 (m, 3H, H-4’’’’’, H-4’, H-4’’’), 5.17 (d, J = 7.9 Hz, 1H, NHCOCH ),
hour at the same temperature, then quenched with Et N, filtered 5.12 (dd, J = 10.4, 7.9 Hz, 1H, H-2’’’’’), 5.01 – 4.95 (m, 3H, H-1, H-1’’,
over Celite and evaporated in vacuo. Crude was purified by flash H-3’’’’’), 4.92 (d, J = 11.1 Hz, 1H, CHHPh), 4.90 – 4.80 (m, 4H, H-3’’’’,
column chromatography (see Supplementary Information). CHHPh, CH Ph), 4.72 – 4.61 (m, 4H, H-1’’’’, CH Ph, H-6’’’’a), 4.60 –
.36 (m, 11H, H-1’’’, H-1’’’’’, H-1’, 4 CH Ph), 4.32 (d, J = 11.9 Hz, 1H,
-Azidoethyl (2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl)-(14)- CHHPh), 4.28 (d, J = 11.9 Hz, 1H, CHHPh), 4.19 (dd, J = 9.4, 8.1 Hz, 1H,
3,6-di-O-acetyl-2-deoxy-2-(2,2,2-trichloroethoxycarbonylamino)- H-3’’), 4.12 – 4.09 (m, 2H, H-6’’’’’a, H-6’’’’’b), 4.09 – 3.90 (m, 6H, H-
6’’’’b, OCHHCH , H-4, H-3, H-4’’, H-2’’’’), 3.90 – 3.86 (m, 1H, H-
5’’’’’), 3.84 – 3.74 (m, 3H, H-6a, H-6’’a, H-4’’’’), 3.74 – 3.63 (m, 4H, H-
6b, H-6’’b, H-3’’’, OCHHCH ), 3.62 – 3.58 (m, 2H, H-5’’’’, H-3’), 3.58
– 3.41 (m, 6H, H-5’, H-5, H-5’’, H-2’, H-2’’’, OCH CHHN ), 3.41 – 3.29
for 1 hour, then the mixture was cooled to −20 °C and NIS (1.5 eq) (d, J = 9.6 Hz, 1H, NHCOCF
3
3
3
3
2
2
4
2
2
[
β-D-glucopyranoside]-(13)-(4-O-acetyl-2,6-di-O-benzyl-β-D-
galactopyranosyl)-(14)-(3,6-di-O-benzyl-2-deoxy-2-
trifluoroacetamido-β-D-glucopyranosyl)-(13)-(4-O-acetyl-2,6-di-
O-benzyl-β-D-galactopyranosyl)-(14)-2-acetamido-3,6-di-O-
2 3
N
2 3
N
2
3
f
benzyl-2-deoxy-β-D-glucopyranoside (56) R = 0.31, cHex/Acetone (m, 5H, H-2, H-6’a, H-6’b, H-6’’’a, H-6’’’b), 3.24 (m, 2H, H-2’’,
This journal is © The Royal Society of Chemistry 20xx
J. Name., 2013, 00, 1-3 | 11
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O rP gl ea na is ce &d Bo i on mo to al e dc juu l sa tr mC haer mg i ins ts ry
Page 12 of 15
ARTICLE
Journal Name
OCH
2
1
2
CHHN
.04 (s, 3H, OCOCH
.89 (s, 3H, NHCOCH
3
), 2.15 (s, 3H, OCOCH
), 2.03 (s, 3H, OCOCH
), 1.49 (s, 3H, NHCOCH
3
), 2.09 – 2.06 (m, 9H, 3 OCOCH
3
), (NHCOCH
3
), 20.8, 20.74, 20.73, 20.69, 20.65, 20.63, 20.61, 20.5 (8
View Article Online
DOI: 10.1039/C8OB03066A
), 1.98 (s, 6H, 2 OCOCH
3
), OCOCH
3
); 19F NMR (376 MHz, CDCl
3
) δ -75.88 (NHCOCF
40: 2447.7546 [M+Na] ; found:
3
+
); HRMS
3
3
13
+
3
3
); C NMR (101 MHz, (ESI ): m/z calcd for C117
H
3
134Cl F
3
N
6
O
3
CDCl ) δ 170.8, 170.5, 170.40, 170.36, 170.12, 170.06, 170.01, 2447.6873.
1
1
1
1
69.95, 169.9, 169.2 (10 COCH ), 157.0 (q, J = 37.3 Hz, NHCOCF ),
3
3
38.9, 138.8, 138.7, 138.4, 138.13, 138.05, 138.0, 137.9 (8 CAr),
28.6, 128.4, 128.32, 128.30, 128.12, 128.08, 128.06, 128.0, 127.9,
27.81, 127.77, 127.64, 127.61, 127.57, 127.5, 127.4, 127.0, 126.8
Conclusions
The synthesis of several diverse lactosamine based building
blocks from lactosamine hydrochloride was successfully
developed. A set of disaccharide blocks bearing different amine
protecting groups (Troc, Phth, TFAc, Ac) was prepared either as
(
3
40 CAr), 115.5 (ad, J = 288.3 Hz, NHCOCF ), 102.4 (C-1’), 102.22 (C-
1
2
’’’), 101.2 (C-1’’’’’), 100.4 (C-1), 100.1 (C-1’’’’), 99.5 (C-1’’), 80.2 (C-
’’’), 79.7 (C-2’), 78.8 (C-3’), 78.0 (C-4’’), 77.2 (C-3’’), 76.6 (C-3), 76.1
(
C-3’’’), 75.9 (C-4), 75.6 (C-4’’’’), 75.1, 75.0, 74.9 (CH
2
Ph, CH
2
Ph, H-5’’,
2
-azidoethyl equipped glycosyl acceptors, or as versatile
H-5), 74.0 (CH
2
Ph), 73.53, 73.49, 73.3, 73.2 (5 CH
2
Ph), 72.8, 72.7 (C-
thioglycoside donors. The availability of a library of lactosamine
based building blocks, all synthesised on a multi-gram scale,
opens the possibility for glycosylation reactivity testing and
constitutes an essential synthetic toolbox for the preparation of
similar structures, e.g. various lactosamine containing N-glycan
fragments. Particularly, the work presents a successful example
of the use of trifluoroacetamide bearing glycosyl donors in the
build-up of LacNAc oligosaccharides, with the best glycosylation
yields (>80%) always obtained for this type of glycosyl donor.
The reactions with trifluoroacetamide donors were always
completely stereoselective (1,2-trans linkage) and no oxazoline
formation was observed. The strategic assembly of the
synthesised set of glycosyl donors and acceptors puts emphasis
on the need to tune acceptor reactivity. The developed high
yielding methodologies for [2+2] glycosylations were applied to
the synthesis of the desired NHTFAc containing
5
(
6
’, C-5’’’), 72.3 (C-5’’’’), 71.9 (C-3’’’’), 70.9 (C-3’’’’’), 70.8 (C-5’’’’’), 69.7
C-4’, C-4’’’), 69.1 (C-2’’’’’), 68.5 (C-6’), 68.3 (OCH CH ), 68.2, 68.0,
7.9 (C-6, C-6’’, C-6’’’), 66.6 (C-4’’’’’), 61.1 (C-6’’’’), 60.9 (C-6’’’’’), 56.7
CH ), 23.6 (NHCOCH ),
); 19F
2
2 3
N
(
C-2’’), 56.4 (C-2), 54.5 (C-2’’’’), 50.6 (OCH
2
2
N
3
3
2
3.1 (NHCOCH
3
), 20.79, 20.75, 20.63, 20.59, 20.5 (8 OCOCH
3
+
NMR (376 MHz, CDCl
for C116
3
) δ -75.95 (NHCOCF
39: 2315.8609 [M+Na] ; found: 2315.8827.
3
); HRMS (ESI ): m/z calcd
+
H
135
F
3
6
N O
2
-Azidoethyl (2,3,4,6-tetra-O-acetyl-β-D-galactopyranosyl)-(14)-
[3,6-di-O-acetyl-2-deoxy-2-(2,2,2-trichloroethoxycarbonylamino)-
β-D-glucopyranoside]-(13)-(4-O-acetyl-2,6-di-O-benzyl-β-D-
galactopyranosyl)-(14)-(2-acetamido-3,6-di-O-benzyl-2-deoxy-
β-D-glucopyranosyl)-(13)-(4-O-acetyl-2,6-di-O-benzyl-β-D-
galactopyranosyl)-(14)-3,6-di-O-benzyl-2-deoxy-2-
trifluoroacetamido-β-D-glucopyranoside (59) R
f
= 0.20, Tol/Acetone
); H NMR (500 MHz, CDCl ) δ 7.44 – 7.16
m, 40H, HAr), 6.81 (d, J = 7.7 Hz, 1H, NHCOCF ), 5.40 – 5.33 (m, 3H,
H-4’’’’’, H-4’, H-4’’’), 5.25 (d, J = 8.0 Hz, 1H, NHCOCH ), 5.11 (dd, J =
0.4, 7.9 Hz, 1H, H-2’’’’’), 5.03 (d, J = 7.9 Hz, 1H, H-1’’), 4.97 (dd, J =
0.4, 3.4 Hz, 1H, H-3’’’’’), 4.91 – 4.84 (m, 3H, H-1, CH Ph), 4.83 – 4.76
Ph), 4.75 – 4.63 (m, 4H, H-3’’’’, CH Ph, CHHCCl ), 4.61 –
.36 (m, 15H, H-6’’’’a, CHHCCl , 4 CH Ph, H-1’’’’, H-1’’’’’, H-1’’’, H-1’,
CH CCl ), 4.35 – 4.27 (m, 2H, CH Ph), 4.13 – 4.08 (m, 2H, H-
’’’’’a, H-6’’’’’b), 4.08 – 3.95 (m, 6H, H-6’’’’b, OCHHCH , H-3, H-4,
H-4’’, H-3’’), 3.87 (at, J = 6.9 Hz, 1H, H-5’’’’’), 3.83 – 3.48 (m, 14H, H-
20
1
8:2; [훼]D = +6.5 (c 1.0, CHCl
3
3
tetrasaccharides
1 and 2 and the work on [2+4] glycosylations
(
3
demonstrates the possibility of further syntheses of larger
LacNAc oligosaccharides in an efficient way. The presence of a
3
1
1
2
-deoxy-2-trifluoroacetamide tag on the prepared structures
2
19
constitutes an important tool to allow for future F NMR
interaction studies with LacNAc binding lectins, e.g. galectins, to
further the knowledge into the characteristics of the binding
interaction, both from a structural and dynamics point-of-view.
(
m, 2H, CH
2
2
3
4
3
2
NHCO
6
2
2
3
2
2 3
N
2
, H-2’’’’, H-6’’a, H-6’’b, H-6a, H-6b, OCHHCH
H-4’’’, H-3’’’, H-3’, H-2’), 3.49 – 3.38 (m, 5H, CH
H-5’’, H-2’’’), 3.38 – 3.25 (m, 5H, H-2’’, OCH
H-6’b), 2.15 (s, 3H, OCOCH ), 2.09 – 2.05 (m, 12H, 4 OCOCH
H, OCOCH ), 2.01 (s, 3H, OCOCH ), 2.00 (s, 3H, OCOCH ), 1.98 (s, 3H,
NHCOCH ) δ 170.5, 170.4, 170.3, 170.1,
); 13C NMR (101 MHz, CDCl
70.03, 169.99, 169.2, 169.94, 169.93 (9 COCH ), 156.7 (ad, J = 37.4
Hz, NHCOCF ), 154.1 (NHCO CH CCl ), 138.8, 138.6, 138.4, 138.3,
38.0, 137.92, 137.89 (8 CAr), 128.7, 128.4, 128.3, 128.22, 128.18,
28.1, 127.9, 127.81, 127.79, 127.73, 127.66, 127.6, 127.54, 127.47,
27.1, 127.0 (40 CAr), 115.6 (ad, J = 286.9 Hz, NHCOCF ), 102.8 (C-1’),
02.1 (C-1’’’), 101.2 (C-1’’’’’), 101.0 (C-1’’’’), 100.3 (C-1’’), 99.0 (C-1),
5.6 (CH CCl ), 80.6 (C-2’’’), 79.4 (C-2’), 77.9 (C-3’), 77.2 (C-3’’), 76.3,
6.1, 76.0 (C-3’’’, C-4’’’’, C-4’’, C-4, C-3), 75.3, 75.0 (C-5, C-5’’, CH Ph,
Ph), 74.3 (CH CCl ), 73.9, 73.6, 73.5, 73.33, 73.26 (6 CH Ph), 73.0,
2
N
3
, H-5’’’’, H-5’’’, H-5, Conflicts of interest
2
CHHN
, H-6’’’a, H-6’’’b,
), 2.02 (s,
3
, H-6’a, H-5’,
There are no conflicts to declare.
2
CHHN
3
3
3
3
3
3
3
Acknowledgements
3
3
We gratefully acknowledge financial support from European
Commission’s GLYCOPHARM ITN project (contract no. 317297)
and Science Foundation Ireland (Grant no. 13/IA/1959).
1
3
3
2
2
3
1
1
1
1
9
7
3
Notes and references
2
3
1
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CH
2
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This journal is © The Royal Society of Chemistry 20xx
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Organic & Biomolecular Chemistry
View Article Online
DOI: 10.1039/C8OB03066A
✓
Important toolbox of lactosamine-based building blocks
[2+2] and [2+4] successful synthetic approaches
✓
✓
High yielding glycosylations with NHCOCF3 donors
NPG = NHTroc, NHCOCF3, NPhth
R = Ac, Bn
NPG = NHTroc, NHCOCF3, NPhth
Assembly
of building blocks
R = Ac, Bn
R = Ac, Bn
Synthesis of fluorine tagged LacNAc
oligomers
19
The synthesis of F containing LacNAc oligomers through the strategic assembly of a small library of
LacNAc/NTFAc disaccharide building blocks