Synthesis of Clostridium bolteae Capsular Polysaccharide Fragments: A Repeating
Disaccharide Unit
by
Jeffrey Davidson
A Thesis
Presented to
The University of Guelph
In partial fulfilment of requirements
for the degree of
Master of Science
in
Chemistry
Guelph, Ontario, Canada
© Jeffrey Davidson, December, 2016
ABSTRACT
SYNTHESIS OF CLOSTRIDIUM BOLTEAE CAPSULAR POLYSACCHARIDE
FRAGMENTS: A REPEATING DISACCHARIDE UNIT
Jeffrey Davidson
Advisor: Dr. France-Isabelle Auzanneau
University of Guelph
Recent studies have shown a link between people with autism who have gastrointestinal
disorders and the bacteria in their gut microflora. Clostridium bolteae has been isolated from
stool samples of children with autism, and a surface capsular polysaccharide has been
determined to have a repeating disaccharide unit comprised of rhamnose and mannose units:
[→3)-α-D-Manp-(1→4)-β-D-Rhap-(1→]. However, D-rhamnose is an extremely rare sugar, and
moreover sequencing of Clostridium bolteae did not show any of the enzymes necessary to
biologically synthesize D-rhamnose. Therefore, this study is to confirm the absolute structure of
this capsular polysaccharide. In this thesis the synthesis of a novel D-quinovose donor is
described as well as its glycosylation with a mannose acceptor. Inversion at O-2 of the quinovose
residue and subsequent glycosylation gave the D-rhamnose trisaccharide. Additionally, progress
towards the L-rhamnose trisaccharide is described. The use of 2,3-O-carbonate or O-2 sulfonyl
donors did not show β-selectivity. Inversion to L-quinovose was accomplished by a Lattrell-Dax
inversion in low yields which should allow for a β-selective glycosylation.
Acknowledgements
I would like to take this opportunity to thank my advisor, Dr. France-Isabelle Auzanneau.
You have given me the opportunity to learn about carbohydrate synthesis and develop many
different skills under your advisement. You have provided constant support and advice
throughout this research project, and I am most thankful.
I would also like to thank Dr. Mario Monteiro for help funding the project as well as
completing much of the background work related to the project.
I would also like to thank my advisory committee members Dr. William Tam and Dr.
Adrian Schwan for the ongoing support and guidance. would also like to thank Dr. Richard
Manderville for representing the graduate faculty, and the chair Dr. Wojciech Gabryelski.
I would also like to thank all the members of the Auzanneau research group, past and
present. I am very grateful for all their help and company in the lab, and the memorable
experiences we had both inside and outside the lab.
iii
Table of Contents
Abstract
List of Schemes
vi
List of Figures
viii
List of Tables
ix
List of General Abbreviations
x
List of Chemical Abbreviations
xii
Chapter 1
Introduction
1
1.1
Introduction
1
1.2
Clostridium bolteae
1
1.2.1
3
1.3
1.4
1.5
1.6
1.7
Biosynthetic pathways of D- and L-rhamnose
Carbohydrate Chemistry Basics
4
1.3.1
Common Sugar Types
7
1.3.2
Glycosylation Reactions
7
Synthesis of Carbohydrates
8
1.4.1
Stannylene Acetals
9
1.4.2
The Anomeric Effect
10
1.4.3
Anomeric Effect in Mannose
12
1.4.4
Identifying Anomeric Configuration
14
1.4.5
Neighbouring Group Participation
15
Glycosyl Donors
16
1.5.1
Thioglycosides
16
1.5.2
Glycosyl Halides
17
1.5.3
Trichloroacetimidates
19
Formation of 1,2-cis mannosides
21
1.6.1
The 4,6-O-Benzylidene acetal
21
1.6.2
The 2,3-O-Carbonate
22
1.6.3
Inversion at O-2
23
Scope of Thesis
24
iv
Chapter 2
Synthesis of D-rhamnose fragment trisaccharide
25
2
Introduction
26
2.1
Synthesis of Mannosyl acceptor 3
27
2.2
Synthesis of Mannosyl donor 4
28
2.3
Synthesis of D-rhamnose unit 5
28
2.3.1
Synthesis of fully protected 14 through acetal pathway
30
2.3.2
Synthesis of disaccharide 18
31
2.3.3
Synthesis of disaccharide acceptor 26
34
2.3.4
Synthesis of trisaccharide 27
36
2.4
Conclusion
Chapter 3
37
Synthesis of L-rhamnose trisaccharide fragment
38
3
Introduction
39
3.1
Glycosylations with the 2, 3-O-carbonate rhamnosyl donor 32
39
3.2
Glycosylation with neighbouring group participation
43
3.2.1
Inversion attempts with benzylated rhamnosyl triflate 39
43
3.2.2
Inversion of benzoyl rhamnosyl triflate 43
46
3.3
Glycosylations with non-participating sulfonyl groups
48
3.4 Conclusion
49
Chapter 4
Future Work
51
4.1
Synthesis of L-trisaccharide 59
52
4.2
Deprotection and NOE analysis
53
4.3
Disaccharide fragments 60 and 61
54
4.3.1
Synthetic strategy for the synthesis of 60
55
4.3.2
Synthetic strategy for the synthesis of 61
56
Chapter 5
Experimental
58
Chapter 6
References
103
v
List of Schemes
Scheme 1: Biosynthetic pathway of L-rhamnose in bacteria cells.
3
Scheme 2: Biosynthetic pathway of D-rhamnose.
4
Scheme 3: Conformations of L and D-Glucopyranose.
6
Scheme 4: Structure of the α- and β-anomers in hexoses.
6
Scheme 5: A general glycosylation reaction.
8
Scheme 6: Glycosylation with participating group at C-2 forming a 1,2-trans glycoside. 15
Scheme 7: Formation of the orthoester and rearrangement to the 1,2-trans glycoside.
15
Scheme 8: Conditions used to prepare thioglycoside donors.
16
Scheme 9: Activation of thioglycoside donors using an electrophilic reagent.
17
Scheme 10: Conditions used to prepare glycosyl halides donors.
18
Scheme 11: Methods to activate glycosyl halides by heavy metals.
19
Scheme 12: Mechanism of the formation of trichloroacetimidate donor.
20
Scheme 13: Mechanism of the activation trichloroacetimidate donors.
20
Scheme 14: Examples of β-mannoside via 4,6-benzylidene acetal.
Taken from Crich and Sun.
22
Scheme 15: Examples of β-mannoside via the 2,3-O-carbonate and insoluble catalysis.
Taken from Crich et al.
23
Scheme 16: Examples of β-mannoside via reductive inversion.
Taken from Wu and Bundle.
24
Scheme 17: Monosaccharide building blocks of trisaccharide 1.
26
Scheme 18: Synthetic steps using to prepare mannosyl acceptor 3.
27
Scheme 19: Synthetic steps using to prepare mannosyl donor 4.
28
Scheme 20: Synthetic steps using to prepare 14 by selective chloroacetylation.
29
Scheme 21: Alternative acetal synthetic pathway to 14 and the synthetic step used
to prepare glycosyl donor 5.
30
Scheme 22: Glycosylation reaction of mannosyl acceptor 3 and glycosyl donor 5.
32
Scheme 23: Hypothesized mechanism for the formation of 24.
35
vi
Scheme 24: Synthetic steps using to prepare disaccharide acceptor 26.
36
Scheme 25: Glycosylation reaction to for trisaccharide 27.
37
Scheme 26: Monosaccharide building blocks being used for the synthesis of 2.
39
Scheme 27: Synthetic steps for the synthesis of bromide 32.
40
Scheme 28: Glycosylation reaction of mannosyl acceptor 3 and rhamnosyl bromide 32. 41
Scheme 29: The β-directing effect of the nitrilium ion.
43
Scheme 30: Synthetic steps to rhamnosyl triflate 39.
44
Scheme 31: Synthetic steps to several rhamnose sulfonates 43, 44 and 45.
47
Scheme 32: Synthetic steps to rhamnosyl trichloroacetimidates 50 and 51.
48
Scheme 33: Glycosylation reaction of mannosyl acceptor 3 and rhamnosyl
sulfonate 44 and 45.
49
Scheme 34: Proposed synthetic steps to trisaccharide 59.
52
Scheme 35: Global deprotection of trisaccharides 28 and 68 giving CPS
fragments 1 and 2.
53
Scheme 36: Proposed synthetic steps to CPS fragment 60.
56
Scheme 37: Proposed synthetic steps to CPS fragment 61.
57
vii
List of Figures
Figure 1: Structure of the Clostridium bolteae CPS disaccharide repeating unit.
2
Figure 2: D-glucose and L-glucose represented in Fischer projection.
5
Figure 3: Structure of L-rhamnose, D-glucose and D-mannose.
7
Figure 4: Formation of the monofluoronated dimer and its selectivity.
Reproduced from Lu et al.
10
Figure 5: The dipole-dipole effect to explain the anomeric effect :
(1) partial cancellation, and (2) overall increase in the dipole moment of the molecule.
11
Figure 6: Explanation of the anomeric effect using the molecular orbital theory and
stereoelectronic stabilization.
12
Figure 7: The dipole-dipole enhancement of mannose-type sugars.
12
Figure 8: The Δ-2 effect in mannose. Newman projection of C2-C1.
Steric repulsion is reduced in α-conformer.
13
Figure 9: Increase of anomeric effect by antibonding-antibonding orbital overlap.
13
Figure 10: The H-1, H-2 dihedral angle in glucose and mannose sugars.
14
Figure 11: Example of a β-glycoside.
21
Figure 12: Structures of the two trisaccharide targets.
24
Figure 13: NOE interactions of Clostridium bolteae CPS. Taken from Pequegnat et al.
54
Figure 14: Target disaccharide Clostridium bolteae CPS fragments 60 and 61.
55
viii
List of Tables
Table 1: Protecting group used in this thesis.
8
Table 2: Glycosylation conditions for the glycosylation reaction of mannosyl
acceptor 3 and glycosyl donor 5.
34
Table 3: Glycosylation reaction conditions of mannosyl acceptor 3 and
rhamnosyl bromide 31.
42
Table 4: Synthetic attempts to invert rhamnosyl triflate 39.
46
Table 5: Synthetic attempts to invert rhamnosyl sulfonates 43, 44 and 45.
47
ix
List of General Abbreviations
Å
Angstroms
aq.
Aqueous
bs
Broad singlet
c
Concentration
°C
Degrees Celsius
13
Carbon Nuclear Magnetic Resonance Spectroscopy
COSY
Proton-Proton Correlation Spectroscopy
CPS
Capsular polysaccharide
δ
Chemical shift
d
Doublet
dd
Doublet of a doublet
ddd
Doublet of a doublet of a doublet
dq
Doublet of a quartet
dt
Doublet of a triplet
equiv
Equivalents
g
Grams
hr
Hours
1
Proton Nuclear Magnetic Resonance Spectroscopy
C NMR
H NMR
HPLC
High-Performance Liquid Chromatography
HRESIMS
High-Resolution Electrosprsay Ionization Mass Spectrometry
HSQC
Heteronucleur Single Quantum Correlation Spectroscopy
Hz
Hertz
J
Coupling constant
JMOD
J-Modulated Spin-Echo Nuclear Magnetic Resonance Spectroscopy
L
Litres
m
Multiplet
mg
Milligrams
x
MHz
Megahertz
min
Minutes
mL
Millilitres
mmol
Millimoles
mol
Moles
MS
Mass Spectrometry
NMR
Nuclear Magnetic Resonance Spectroscopy
q
Quartet
quin
Quintet
quant.
Quantitative
Rf
Retardation factor
RP
Reverse-Arase
RT
Room temperature
s
Singlet
sat’d
Saturated
SN1
Nucleophilic Substitution Order 1
SN2
Nucleophilic Substitution Order 2
t
Triplet
td
Triplet of a doublet
TLC
Thin-Layer Chromatography
μL
Microlitres
xi
List of Chemical Abbreviations
Ac
Acetyl
All
Allyl
Bn
Benzyl
Bu
Butyl
Bz
Benzoyl
ClAc
Chloroacetyl
CSA
10-Camphorsulfonic Acid
DBU
1,8-Diazabicyclo[5.4.0]undec-7-ene
DMAPA
3-(Dimethylamino)-1-propylamine
DMF
N,N-Dimethylformamide
DMP
2,2-Dimethylpropane
DMSO
Dimethylsulfoxide
Et
Ethyl
Glc
Glucose
Lev
Levulinoyl
Man
Mannose
Me
Methyl
Ms
Methanesulfonyl
NIS
N-Iodosuccinimide
Ar
Phenyl
Piv
Pivaloyl
py
Pyridine
Rha
Rhamnose
SiO3
Silicate
TBAI
Tetrabutylammonium iodide
Tf
Trifluoromethanesulfonyl
TFA
Trifluoroacetic acid
xii
THF
Tetrahydrofuran
TMS
Trimethylsilyl
Ts
p-toluenesulfonyl
xiii
CHAPTER 1 – Introduction and Background
1.1
Introduction
Carbohydrates are the most abundant class of natural products found in living organisms.
Carbohydrates act as an energy source for both plants and animals and can be used in a structural
role.1 Carbohydrates are also abundant on the surface of cells and bacteria with bacteria often
being enveloped by a rigid polysaccharide layer known as the capsular. The capsular is coated
with various polysaccharides, which are referred to as capsular polysaccharides (CPS).1 Capsular
polysaccharides create an outside mucus-like layer protecting the cell from the external
environment and from a non-specific host immune responses.2 Additionally, the CPS layer is
involved in interactions between the bacteria and the external environment, such as signalling.
Capsular polysaccharides are often very unique and can be specific to a particular
bacterium.2 Additionally, CPS can be immunogenic which would result in an immune response
from the host. Due to the uniqueness of bacterial capsular polysaccharides, any invoked immune
response would be very specific to the particular bacteria, making these polysaccharides an
excellent target for glycoconjugate vaccines.
1.2
Clostridium bolteae
The Clostridium genus is comprised of Gram-positive bacteria which several are known
for producing neurotoxins.3,4 Clostridium bacteria are anaerobic and often reside within the gut
mircoflora.3 These bacteria are known to produce short chain fatty acids (SCFAs) which can
alter the contraction rate of the gastrointestinal (GI) tract.5 Changes to the intestinal contraction
rate increases the likelihood of GI tract disorders such as diarrhea. Children with autism are
commonly afflicted with GI disorders and recently a link has been established between GI
disorders and the bacteria in their gut microflora.5,6,7 Interestingly, autistic children have
1
significantly different gut microflora compared to their non-autistic counterparts.6 In 2003, Song
et al. were able to isolate a bacterium strain, Clostridium bolteae, from the stool samples of
autistic children suffering from various GI disorders.8 The capsular of strains 16351 and 14578
had their structure analyzed by Pequegnat et al. and was shown to express an unique capsular
polysaccharide.9 In immunogenic testing this capsular polysaccharide invoked an immune
response with rabbits. Immunodot blots of the rabbit serum indicated strong interactions of the
IgG antibodies with the CPS even at a dilution of 1:1000.9 The strong immunogenic response
observed in rabbits suggests that this CPS is a good target for the development of an antiClostridium bolteae vaccine.
Figure 1: Structure of the Clostridium bolteae CPS disaccharide repeating unit.
The structure of a Clostridium bolteae capsular polysaccharide was identified through
various different analytical techniques. The glycosidic linkages were hydrolyzed in acidic
conditions to their individual monosaccharide units. Upon acetylation the sugars were subjected
to GC-MS and through comparison to known analytical samples, each sugar and its
configuration was assigned.9 The capsular polysaccharide has been identified as a repeating
disaccharide unit, comprised of D-rhamnose (6-deoxymannose) unit linked to a D-mannose via a
1,3-β-glycosylic linkage and repeated through 1,4-α-glycosylic linkage (Figure 1).9 Additional
NOE-NMR experiments and molecular modelling further support this structural assignment.
2
1.2.1 Biosynthetic pathways of D- and L-rhamnose
The deoxy-sugar rhamnose exists as two enantiomers, D-rhamnose and L-rhamnose. Lrhamnose is the more common of the two and is present in both plants and bacteria while Drhamnose is quite rare and found exclusively in bacteria.10,11 The biosynthesis of the two
rhamnose sugars differs significantly. L-rhamnose’s biosynthetic pathway has been well
established, with two different pathways for plant and bacteria cells.10a The bacterial synthetic
pathway begins with dTDP-D-glucose and 3 enzymes is convert it to dTDP-L-rhamnose
(Scheme 1).10a The enzymes dTDP-Glc 4,6-dehydratase (rmlB) and 3,5-epimerase (rmlC)
convert dTDP-D-glucose to the intermediate dTDP-4-keto-6-deoxy-L-mannose and dTDP-4ketorhamnose reductase (rmlD) finishes the conversion to dTDP-L-rhamnose.10a
dTDP-D-glucose
H2O
rmlB
dTDP-4-keto-6-deoxy-D-glucose
rmlC
dTDP-4-keto-L-rhamnose
NADPH
rmlD
dTDP-L-rhamnose
Scheme 1: Biosynthetic pathway of L-rhamnose in bacteria cells.10a
D-rhamnose follows a similar biological pathway to several different deoxyhexoses
sugars, such as L-fucose and 6-deoxy-D-talose.11 These deoxy-sugars all have the intermediate
GDP-4-keto-6-deoxy-D-mannose as a common branching point. GDP-4-keto-6-deoxy-D3
mannose is formed from the catalysis of GDP-D-mannose by the enzyme GDP-D-mannose 4,6dehydratase (GMD).11 From the branching point, a D-mannose reductase enzyme (RMD) that
targets the 4-keto group converts the sugar into GDP-D-rhamnose.
GDP-α-D-mannose
H2O
GMD
GDP-4-keto-6-deoxy-D-mannose
RMD
GDP-D-rhamnose
Scheme 2: Biosynthetic pathway of D-rhamnose.11
Interestingly, strain 14578 has since had its genome sequenced. Enzymes rmlB, rmlC and
rmlD have all been identified.12 These are the enzymes required for the biological synthesis of Lrhamnose, while none of the enzymes required for to biologically synthesize D-rhamnose have
yet to be observed.12 Additionally, other L-rhamnose enzymes, L-rhamnose mutarotase and Lrhamnose isomerase, have been sequenced.12 However, there is no evidence to suggest that these
L-rhamnose enzymes are directly linked to this exact capsular polysaccharide and not elsewhere
within the bacterium. In addition, the lack of D-rhamnose enzymes within the bacterium may not
be conclusive as those enzymes may have yet to be sequenced. Therefore, the goal of this
research is to confirm the absolute configuration of this CPS.
4
1.3
Carbohydrate Chemistry Basics
Monosaccharides are the basic building blocks of carbohydrates. Numbering of
carbohydrates begins with the highest oxidized carbon. Each monosaccharide exists as two
enantiomers and to distinguish between the two the prefixes D and L are used. When represented
in a Fischer projection D-sugars have the hydroxyl group of their highest numbered stereocenter
on the right, where L-sugars have this hydroxyl on the left. These letters refer to the most distant
stereocenter from the carbonyl group, which in hexoses is C-5 (Figure 2).
With hexoses, these sugars are much more stable as a hemiacetal and cyclize to form 6
membered rings. D-hexoses generally adopt a 4C1 chair conformation allowing for the most
substituents to be equatorial on the ring while L-hexoses adopt the 1C4 conformation (Scheme
3).1 The 6 membered ring results from intramolecular nucleophilic attack of the O-5 hydroxyl
group on the aldehyde group.
Figure 2: D-glucose and L-glucose represented in Fischer projection.
Cyclization causes a new stereocenter to be formed at C-1, creating two different
configurations as the aldehyde is converted to a hemiacetal. If the newly formed hydroxyl group
forms on the same side as the C-H bond of the highest numbered stereocenter (which in a sixmembered ring is C-5) then the α-anomer is formed, while the β-anomer has its anomeric
hydroxyl group on the opposite side as this C-H bond (Scheme 4).
5
4
C1
D-Glucopyranose
4
C1
L-Glucopyranose
1
C4
D-Glucopyranose
1
C4
L-Glucopyranose
Scheme 3: Conformations of L and D-Glucopyranose.
α-anomer
β-anomer
Scheme 4: Structure of the α- and β-anomers in hexoses.
6
1.3.1 Common Sugar Types
There are various types of monosaccharide with some common ones being glucose and
mannose. Glucose is the most common sugar, having all hydroxyl groups equatorial on the ring.
Mannose is the C-2 epimer of glucose, having O-2 in the axial position.1 In this thesis
monosaccharides D-glucose, D-mannose and L-rhamnose (6-deoxy-L-mannose) are used.
L-rhamnose
D-glucose
D-mannose
Figure 3: Structure of L-rhamnose, D-glucose and D-mannose.
1.3.2 Glycosylation Reactions
A glycosylation occurs when a sugar is bound covalently to another sugar or protein
through the formation of a glycosidic bond. In a glycosylation, two major species are present, the
glycosyl donor and the glycosyl acceptor. The glycosyl donor is fully protected and equipped
with a good leaving group at the anomeric position. When the leaving group is activated,
generally with a Lewis acid, an oxocarbenium ion intermediate is formed. The anomeric carbon
of the oxocarbenium ion is sp2 hybridized, allowing for a nucleophilic attack from a hydroxyl
group of the glycosyl acceptor creating a glycoside (Scheme 5).13
7
Scheme 5: A general glycosylation reaction.
1.4 Synthesis of Carbohydrates
Protecting group manipulation is a common practice in organic synthesis. In synthetic
carbohydrate chemistry protecting groups are employed to ensure proper stereoselectivity and
regioselectivity. As previously mentioned glycosyl donors are fully protected with a good
leaving group at the anomeric centre, while glycosyl acceptors are fully protected besides the
nucleophilic OH to ensure the proper regioselectivity. Many factors of protecting groups can
influence the stereoselectivity of a reaction. A few factors that may influence a reaction are the
size, the electronic effects or the rigidness of the protecting groups.
The ideal protecting group is one that is able to be introduced in mild conditions and is
easily removed. Many protecting groups can be cleaved exclusively using specific conditions
allowing for selective removal later in the synthesis.1,13 A wide range of protecting groups have
been used in this thesis, which are outlined in Table 1.
8
Table 1. Protecting group used in this thesis
Protecting Group
Method of Addition
Method of Removal
O-Acetate –OAc
Ac2O/pyridine
HCl/MeOH
O-Chloroacetate – OAcCl
ClAcCl/pyridine
Thiourea
O-Benzoate – OBz
BzCl/pyridine
Na/NH3(l)
O-Levulinoate – OLev
Lev2O/pyridine/DMAP
H2NNH2•HOAc
di-O-Carbonate
Arosgene/pyridine
Na/NH3(l)
O-Benzyl – OBn
BnBr/NaH
Na/NH3(l)
O-Allyl –OAll
AllBr/NaH
1. RhCl(PPh3)3, DABCO
2. HgCl2, HgO
di-O-Isopropylidene
Acetone/p-TsOH
80% AcOH
Ester Type:
Ether Type
1.4.1 Stannylene Acetals
Stannylene acetals were first reported in 1974 and since have seen widespread use in
carbohydrate synthesis.14 Stannylene acetals are commonly formed from diols with alkyl tin
oxides, such as dibutyltin oxide, in toluene or benzene with the azeotropic removal of water.
Stannylene acetals have risen in popularity due to their ability to react with electrophiles giving
the monosubstituted product with high regioselectivity.15 Stannylene acetals are typically used
for ether protection of alcohols, and in a cis-diol like mannose selective protection of the
equatorial alcohol is achieved. Stannylene acetals can also be used in acetyl protection of
9
alcohols, however, usually with a much lower regioselectivity.
Improvements to the reactions times and regioselectivity can be accomplished upon the
addition of a nucleophile, like cesium fluoride or tetrabutylammonium iodide.16,17 The exact
mechanism for the increase in selectivity is not fully understood, but it is believed to be the result
of the nucleophile coordinating to the tin. The coordination to the nucleophile may cause the tin
to aggregate into dimers and higher oligomers. However, computational analysis suggests that
these dimers prefer to be only coordinated to a single nucleophile. Calculations showed that
difluoridated dimers are between to be 209 to 278 kJ/mol less stable than their monofluoridated
dimer counterparts.16 Mono substitution of the dimers creates a unique highly nucleophilic
oxygen allowing for high stereoselectivity (Figure 4).
Figure 4: Formation of the monofluorodated dimer and its selectivity. Reproduced from Lu et al.17
1.4.2 The Anomeric Effect
Substituents can exist either equatorially or axial on a ring. When a substituent is
positioned axially it experiences greater steric hindrance making its equatorial counterparts more
energetically favoured. However, at the anomeric position this rule does not fully apply and this
exception is referred to as the anomeric effect. The anomeric effect refers to the preferred
bonding arrangments of electronegative substituents at the anomeric centre.18 In 6-membered
10
rings, the axial orientation for electronegative substituents is usually more stable and is preferred
in most cases. The anomeric effect can be explained by two theories. The first theory is the
dipole-dipole stabilization.19 When the electronegative substituent is in the α-configuration the
effective dipole of the substituent partially cancels the effective dipole of the endocyclic oxygen,
if the substituent is in the β-configuration the effective dipoles sum together increasing the
overall net dipole moment (Figure 5).19 By partially cancelling the effective dipole, the overall
energy of the sugar is lowered leading to the more stable α-anomer.
α-anomer
β-anomer
Figure 5: The dipole-dipole effect to explain the anomeric effect : (1) partial cancellation, and
(2) overall increase in the dipole moment of the molecule.
The second theory that is used to explain the anomeric effect involves stereoelectronic
stabilization.20 When the electronegative substituent is in the α-configuration, this allows for the
alignment of the non-bonding HOMO of the endocyclic oxygen and the anti-bonding LUMO of
the carbon-oxygen bond. Due to this alignment the HOMO is able to donate into the LUMO
reducing the overall energy of the system and stabilizing the sugar (Figure 6).13,20 When the
substituent is in the β-configuration, this alignment is not possible and the lowering of energy
does not occur.
11
HOMO ny-orbital
LUMO
*
σ C
LUMO σ* C
HOMO
ny
Figure 6: Explanation of the anomeric effect using the molecular orbital theory and
stereoelectronic stabilization.
1.4.3 Anomeric Effect in Mannose
In mannose type sugars, in which the O-2 is axial, the α-anomer is more favoured than
other sugar types. This preference for the α-anomer can be explained by multiple theories. One
theory is an extension of the anomeric effect, with the further stabilization of dipoles. The axial
O-2 exerts an effective dipole in a similar direction of the ring oxygen, and adds to the overall
effective dipole, which is greater when compared to other sugars (Figure 7). Therefore, the αconfiguration is much greatly preferred as it partially cancels this stronger dipole moment.
α-anomer
β-anomer
Figure 7: The dipole-dipole enhancement of mannose-type sugars
The second theory is due to the Δ-2 effect. Axial substituents create elements of
instability in pyranose rings, and this effect is strongest at C-2. When mannose-type sugars are in
the β-configuration, the endocyclic oxygen and the bulkier hydroxyl groups are in close
proximity to each other.21 When observed through a Newman projection, the β-configuration has
12
a small dihedral angle between its the three oxygens, increasing steric hindrance (Figure 8).
However, when in the α-configuration, the dihedral angle of the oxygens is much greater,
reducing steric hindrance, and lowering the sugar’s energy state.
α-anomer
β-anomer
Figure 8: The Δ-2 effect in mannose. Newman projection of C2-C1. Steric repulsion is reduced
in α-conformer.
The prevalence of the anomeric effect in mannose-type sugars can also be explained due
to stereoelectronic effects.22 When O-2 is axial, a secondary overlap between the antibonding
orbitals of C1-O and C2-O is present (Figure 9). This is in addition to the previously mentioned
overlap from the endocyclic oxygen, which furthers stabilized the α-conformer.
Figure 9: Increase of anomeric effect by antibonding-antibonding orbital overlap
1.4.4 Identifying Anomeric Configuration:
Determination of the anomeric configuration is a necessity. Determination of the
anomeric configuration in glucose-type sugars can be achieved through by the 3JHH coupling
constant.23 Depending on which anomer is present, the dihedral angle with H-2 differs and based
13
on the Karplus curve the anomer can be determined by the 3JHH coupling constant.23 When in the
α-anomer, the dihedral angle between the two protons is approximately 60o resulting in a
coupling constant around 3-4 Hz. When the dihedral angle close to 180o, as in the β-anomer, the
coupling constant is 7-9 Hz. However, in mannose type sugars this method cannot be used. If H2 is equatorial regardless of which anomer the dihedral angle is similar, preventing the
determination from the 3JHH coupling constant. Instead the 1JCH coupling constant is used. As
reported by Bock and Pederson, the 1JCH coupling constant differs between the two anomers.24 βanomers have a 1JCH coupling constant of approximately 160 Hz compared to the α-anomers
which have a coupling constant of around 170 Hz.
α-anomer
β-anomer
α-anomer
β-anomer
Figure 10: The H-1, H-2 dihedral angle in glucose and mannose sugars.
1.4.5 Neighbouring Group Participation
When an amide or ester group is located at C-2, participation from this group can occur.
During a glycosylation, such participation allows stereoselective control which directs what
anomer is formed.1,13 After the activation of the anomeric carbon to form the oxocarbenium ion,
14
an acetyl group for example, can act as an internal nucleophile and attack the anomeric carbon,
forming a 1,2-cis acetoxonium ion. This inhibits the ROH nucleophile from attacking from that
side and instead it forms a 1,2-trans glycoside. In glucose, this involves the acetyl group binding
to the anomeric carbon axially and the ROH nucleophile can only attack in the equatorially plane
forming the β-anomer (Scheme 6).13
Scheme 6: Glycosylation with participating group at C-2 forming a 1,2-trans glycoside.
A potential risk in using any participating group is the formation of an orthoester.1,13
Once the acetoxonium ion immediate is formed the electrophilic carbon of the participating
group can be attacked by the nucleophile instead of the anomeric carbon. However, orthoesters
are acid labile and potentially can rearrange to the 1,2-trans glycoside in acidic conditions.1,13
Scheme 7: Formation of the orthoester and rearrangement to the 1,2-trans glycoside.
15
1.5
Glycosyl Donors
Glycosyl donors have a good leaving group at the anomeric centre, which can be
activated to form the oxocarbenium ion. Activation is generally done by a Lewis acid. There is a
variety of types of glycosyl donors and various methods of activation. The glycosyl donors used
in this thesis will be discussed.
1.5.1
Thioglycosides:
Thioglycoside donors have become popular in carbohydrate synthesis due to their ease of
introduction as well as their high chemical stability and versatility. Originally used back in the
early 1900’s by Fischer and Delbruck, thioglycosides are often prepared from an anomeric
acetate with a Lewis acid, usually BF3•OEt2.25 Activation of the acetate leads to the formation of
an oxocarbenium ion, and allows for a nucleophilic attack by a thiol acceptor.
Scheme 8: Conditions used to prepare thioglycoside donors.
The activation of thioglycosides are done using a thiophilic reagent to facilitate the
removal of the anomeric group.1,13 Soft electrophiles, such as X+, can be attacked by the sulfur’s
lone pairs creating a sulfonium intermediate, which is an excellent leaving group and upon
explusion creates the oxocarbenium ion. Subsequent attack of a nucleophile will result in the
formation of a glycosidic bond. Thioglycosides can be easily transformed into other various
glycosyl donors, such as anomeric sulfoxides or anomeric halides.
16
Scheme 9: Activation of thioglycoside donors using an electrophilic reagent.
1.5.2 Glycosyl Halides
Glycosyl halides were originally reported in 1901 by Koenigs and Knörr.26 Glycosyl
halides most common are glycosyl chlorides or bromides, however, glycosyl fluorides have also
been reported.27 Glycosyl halides are used due to their ease of preparation and their high
reactivity. However, these halides are prone to degradation, therefore they must be used
immediately after preparation.
Preparation of glycosyl halides is usually accomplished through one of two methods. One
method is from an anomeric acetate.13 The anomeric acetate can be protonated by a strong acid
(HX), causing the departure of the acetate group and the formation of the oxocarbenium ion. The
X- attacks the anomeric carbon forming the glycosyl halide. Glycosyl halides exist exclusively as
α-anomer due increased stability brought on by the anomeric effect. Another method to form the
glycosyl halide is through a thioglycoside.13 Thioglycosides can be directly converted to the
halide by elemental halogens, typically elemental bromine. The lone pair on sulfur attack the
bromine creating the sulfonium ion. This can be subsequently displaced by the bromide creating
the glycosyl halide.
17
Scheme 10: Conditions used to prepare glycosyl halides donors.
Activation of these donors is usually achieved with soluble heavy metal salts, such a
silver or mercury salts.26,28 Complexing of the glycosyl halide to a soluble metal activates the
donor resulting in the departure of the halide and the formation of the oxocarbenium and the
precipitation of an insoluble salt. Activation of glycosyl halides can also be achieved with
insoluble silver catalysts, such as silver carbonate or silver oxide.29 With insoluble salts the
glycosyl halide deposits on the surface of the silver forming a weak complex. This complex is
susceptible to nucleophilic attack, however, the complexation blocks the α-face leading to the βglycoside through a SN2 type mechanism. Activation by an insoluble catalyst is slower than
soluble salts thus making reactions with poorer nucleophiles difficult.
18
Scheme 11: Methods to activate glycosyl halides by heavy metals.
1.5.3 Trichloroacetimidates
Trichloroacetimidate donors have become the most popular donor type in recent years.
Originally reported by Schmidt and Michel in 1980 trichloroacetimidates have risen in popularity
due to their ease of introduction and activation.30 From the hemiacetal, the trichloroacetimidates
can easily be formed through base-catalysis and trichloroacetonitrile. Deprotonation of the
hemiacetal creates an alkoxide anion. The anion can attack the electrophilic carbon of
trichloroacetonitrile, creating a nitrogen anion which is protonated by the conjugate acid to give
the trichloroacetimidate donor and regenerating the base.13
19
Scheme 12: Mechanism of the formation of trichloroacetimidate donor.
Trichloroacetimidates can be activated by catalytic amounts of a Lewis acids, such as
TMSOTf and BF3•OEt2. The high reactivity of trichloroacetimidate donors allows for activation
even at low temperatures.
Scheme 13: Mechanism of the activation trichloroacetimidate donors.
20
1.6
Formation of 1,2-cis mannosides
Synthesis of 1,2-cis mannosides are the hardest glycosidic bond to form (Figure 11).29, 31
The anomeric effect favours the formation of an α-linkage, which is even more pronounced in
mannose-type sugars. Neighbouring group participation cannot be used as it would lead to the
formation of a α-glycoside. Additionally, as previously mentioned steric effects of the axial O-2
group limits access for the nucleophile to attack. There have been several method developed to
form β-mannosides with some of the common methods being discussed in this thesis.
Figure 11: Example of a β-glycoside.
1.6.1 The 4,6-O-Benzylidene acetal
One of the most common and well known methods to create β-mannosides employs the
4,6-benzylidene acetal. This method originally developed by David Crich gives high βselectivity for a variety of donors and acceptors.32 The benzylidene acetal is known to be
torsionally disarming, lowering the reactivity of the donor while locking the conformation.
Additionally, a triflate anion is used as it forms an triflate intermediate, with the typical reaction
being between a thioglycoside and triflic anhydride. The triflate anion coordinates to the
oxocarbenium ion and due to its high electron withdrawing effect the triflate ion forms as the αanomer, which gets sequentially displaced by the nucleophile in an SN2 type substitution.
21
Scheme 14: Examples of β-mannoside via 4,6-benzylidene acetal. Taken from Crich and Sun.32b
1.6.2 The 2,3-O-Carbonate
In rhamnose (6-deoxymannose) the use of the 4,6-benzylidene acetal is impossible. Other
methods have been developed that work similarly though. The most common method being the
use of the 2,3-O-carbonate protecting group with an insoluble silver catalyst. 31, 33 The effect of
the carbonate was studied by Crich et al. in 2003 and the β-directing nature of the carbonate was
explained through a combination of factors.34 The electron withdrawing effect of the carbonate
group reduces the reactivity of the anomeric bromide. This results in the destabilization of a
oxocarbenium ion making the expulsion of the bromide less favoured and allowing for an SN2
type reaction to occur. This theory is supported by other similar experiments with a 2,3cyclohexylidene.35 The 2,3-cyclohexylidene does not have the same withdrawing effect as the
carbonate and has little to no β-selectivity. Additionally, the insolubility of catalyst is also
important for β-selectivity. The insoluble nature of the catalyst limits the rate of deposition on
the surface by the halide, slowing the formation of the oxocarbenium ion.34 The deposition onto
22
the silver also shields the α-face making a nucleophlic attack from the β-face more favoured.
However, due to the lower reactivity of the glycosyl donor, generally weaker
nucleophiles will not work.29,31
Scheme 15: Examples of β-mannoside via the 2,3-O-carbonate and insoluble catalysis. Taken
from Crich et al.34
1.6.3 Inversion at O-2
Due to the difficulties of forming β-mannosides, methods involving the inversion at C-2
after glycosylation have been developed. This technique has been around for decades and begins
with a equatorial O-2 sugar such as glucose.36,37 Formation of the β-glycoside is achieved
through neighbouring group participation and afterwards O-2 undergoes an oxidation then
reduction with a bulky reducing agent, such as L-selectride®, to produce an axial O-2.38 This
method is highly β-selective, however, is quite inefficient as it requires multiple steps after
glycosylation.
23
Scheme 16: Examples of β-mannoside via reductive inversion. Taken from Wu and Bundle.38
1.7
Scope of Thesis
The goal of the chemistry presented this thesis is to synthesize the two trisaccharide
fragments shown in Figure 12. These fragments contain either a D-rhamnose or L-rhamnose unit
and will be used to confirm the structure of the Clostridium bolteae capsular polysaccharide.
This thesis will contain the synthesis of four monosaccharide building blocks through various
protecting group manipulations.
Figure 12: Structures of the two trisaccharide targets.
24
Chapter 2
Synthesis of D-rhamnose fragment trisaccharide
25
2
Introduction
In order to investigate the absolute configuration of the Clostridium bolteae capsular
polysaccharide we proposed the synthesis of two trisaccharides. These trisaccharides differ in
regards of the absolute configuration of the rhamnose unit. This chapter will focus on the
synthesis of the D-rhamnose trisaccharide 1. Trisaccharide 1 was synthesized from three
monosaccharide building blocks which are shown in scheme 17. The mannose acceptor 3 and
mannose donor 4 have both been previously reported and were synthesized with only mild
alterations to their reported procedures.39,40 Synthesis of a new rhamnose donor 5, will be
described in this chapter. The difference between this donor and previously reported donor is the
chloroacetyl protecting group at the OH-2 position. The chloroacetyl group was chosen as it
allows for selective deprotection in the presence of other acetyl groups.
Scheme 17: Monosaccharide building blocks of trisaccharide 1.
26
2.1
Synthesis of mannosyl acceptor 3
Synthesis of the known mannose acceptor began with commercially available methyl-α-
D-mannose and followed the procedure of Bundle and Eichler with little modification.40,41
Formation of the 2,3-O-isopropylidene was accomplished with acetone and 2,2dimethoxypropane in acidic conditions. Initially both the 4,6-O and 2,3-O acetals were formed,
however, upon the addition of water the more labile 4,6-O-isopropylidene ring was hydrolyzed
giving the 2,3-O-mannose ketal 6 in a 61% yield. Protection of O-4 and O-6 occurred
simultaneously by benzylation with benzyl bromide and sodium hydride to give 7 in 70% yield.
Hydrolysis of the isopropylidene occurred in 80% acetic acid at 80 oC to give the diol 8 in 83%
yield. Temporary formation of an orthobenzoate was achieved using trimethylorthobenzoate in
acetonitrile. This differed from the reported procedure as they used triethylorthobenzoate in
DMF. The orthobenzoate was rearranged with acetic acid to give the O-2 benzoate and the
mannose acceptor 3 in 80% yield which is about a 20% increase over the reported yield.40
Scheme 18: Synthetic steps using to prepare mannosyl acceptor 3.
27
2.2
Synthesis of Mannosyl donor 4
The synthesis of the mannose donor began from commercially available peracetylated D-
mannose, which was subjected to a glycosylation with thiophenol and boron trifluoride diethyl
etherate, producing the thioglycoside 9 in 74% yield. The sugar was deacetylated under Zèmplen
conditions, which was followed by benzoylation giving the thioglycoside donor 4 in 89% yield.39
Benzoyl protecting groups were used in hopes to limit the formation of the orthoester.
Additionally, the benzoyl groups allow us to selectively deprotect acetyl protecting groups,
which is needed in the synthesis of the L-trisaccharide.
Scheme 19: Synthetic steps using to prepare mannosyl donor 4.
2.3 Synthesis of D-rhamnose unit 5
Due to difficulties in forming β-mannosides the synthetic scheme involved the formation
of the β-linkage via neighbouring participation followed by an inversion afterwards. Due to the
high costs of D-rhamnose and D-quinovose, synthesis of the D-donor began with commercially
available diacetone-D-glucose. The free O-3 alcohol was selectively protected using benzyl
bromide and NaH with catalytic amounts of tetrabutylammonium iodide, which was followed by
the hydrolysis of the more labile primary acetal to give the known diol 10 in 87% yield over the
two steps.42 Conversion of the primary alcohol to a tosylate was accomplished at low
temperatures with pyridine and catalytic amounts of DMAP. When cooled to only 0 oC
formation of the O-5, O-6 ditosylate was observed anywhere from 15-25% of 11.43 However,
28
when the reaction was cooled to -20 oC and tosyl chloride was added in 0.5 equivalent additions
which occurred over several days, the primary tosylate was exclusively formed in high yields.
Reduction of the tosylate to give the 6-deoxy sugar was done with LiAlH4.44 The reduction of the
tosylate goes through a 5, 6 oxirane intermediate, which weaker reducing agents like NaBH4
have difficulties reducing. NaBH4 requires high temperatures over long periods of time to reduce
the oxirane, while LiAlH4 accomplished it at room temperature with better yields in under an
hour. Hydrolysis of the 1,2 isopropylidene in 80% acetic acid requirs heat and gave the known
hemiacetal 12 in 71% yield from the tosylate.44 From the hemiacetal two different pathways
were taken to get to donor 5. Initially, synthesis was achieved by selective chloroacetylation. The
reactivity of the hydroxyl group of 12 go from O-1, O-2, O-4, allowing for selective
chloroacetylation to occur.45,46 At -20 oC, O-1 and O-2 were able to be preferentially
chloroacetylated. However, this reaction did not have the highest selectivity and a wide range of
products were formed and resulted in poor yields. Acetylation of O-4 using pyridine and acetic
anhydride gave the protected sugar 14 in 71% yield. The anomeric chloroacetate was labile and
would degrade in these conditions, requiring for the reaction to run for no longer than 15
minutes.
Scheme 20: Synthetic steps using to prepare 14 by selective chloroacetylation.
29
2.3.1 Synthesis of fully protected 14 through acetal pathway
The difficulties in synthesizing 14 led us to consider other synthetic pathways to the
protected sugar 14. According to Jiang and Schmidt, the 1,2 isopropylidene pyranose can be
formed under kinetic conditions, which would allow for the selective protection of O-4.47 When
hemiacetal 12 was diluted in THF with 2,2-dimethoxypropane the kinetic pyranose
isopropylidene was formed in 85% yield. The acetal 15 had O-4 acetylated with acetic anhydride
and pyridine to give 16 in 95%. Hydrolysis of the isopropylidene using the previous conditions
gave the hemiacetal in 78% yield. Chloroacetylation of the diol gave 14 in 93% yields. Between
the two synthetic pathways to 14 the favoured route was through the acetal 15. Although, this
method did require additional steps, it allowed for a much higher selectivity and purification was
much easier. Furthermore, the overall yield between the two routes are similar with both routes
having an overall yield between 40-45%. Selective deacetylation of the anomeric acetate using
dimethylaminopropylamine gave the hemiacetal and was converted to the trichloroacetimidate 5
by trichloroacetonitrile and 1, 8-diazabicyclo[5.4.0]undec-7-ene in a 68% yield over the two
steps.
Scheme 21: Alternative acetal synthetic pathway to 14 and the synthetic step used to prepare
glycosyl donor 5.
30
2.3.2 Synthesis of disaccharide 18.
Synthesis of disaccharide 18 proved to be quite difficult. The challenges arose from both
the high reactivity of the donor, and also the unwanted formation of orthoester 19 was quite
common. The orthoester was identified by 13C and 1H NMR analysis. Evidence of the orthoester
can be observed in the JH-1, H-2 coupling constant. The β-glycosidic bond in disaccharide 18 had a
JH-1, H-2 = 7.9 Hz, while orthoester 19 had a JH-1, H-2 = 5.3 Hz. This is expected as the Karplus
curve predicts that 19 should have lower JH-1, H-2. Additionally, due to the increased electron
withdrawing effect of the orthoester a downfield shift of H-1 of the glucose unit is expected.
This is observed as the H-1 of the glucose residue is at 5.77 ppm in 19, compared to 4.61 ppm in
the disaccharide. It is also expected that H-2 of the glucose residue will shift upfield in 19 which
occurred as H-2 in the orthoester was at 4.55 ppm compared to 5.01 ppm in 18. The CH2Cl
protons of chloroacetyl group split and had a downfield shift in the orthoester to 3.83 ppm and
3.66 ppm as opposed to 3.28 ppm in the disaccharide. In the 13C NMR spectrum the CH2Cl of
the chloroacetyl group had a noticeable downfield shift to 44.3 from 40.2 ppm in the
disaccharide. Further confirmation of the formation of the orthoester was the upfield shift of 4o
chloroacetyl peak. Typically, the 4o chloroacetyl 13C resonance is observed around 169 ppm
while in the orthoester this peak was observed at 119.1 ppm.
Various conditions were attempted for the synthesis of 18 which are summarized in table
2. Donor 5 was able to be activated with catalytic amounts of TMSOTf even at low temperature,
often leading to its degradation. However, when the temperature was lowered or sieves were
used in the reactions, the formation of the orthoester was favored. This was extremely
pronounced when a high amounts of 4Å sieves (200 mg/mL) was used, it resulted in the
formation of the orthoester in good yields (entry 2). Additionally, whenever the donor was used
31
Scheme 22: Glycosylation reaction of mannosyl acceptor 3 and glycosyl donor 5.
in excess the formation of the orthoester was favoured (entry 3). It is hypothesized that when the
donor was in excess the kinetic orthoester product was formed, however, when the acceptor was
in excess it allowed for the thermodynamically favoured disaccharide instead. Furthermore,
when the weaker activator BF3•OEt2 was used at greater equivalents the orthoester was still
formed (entry 4). Attempts to rearrange the orthoester to the disaccharide with additional
equivalents of BF3•OEt2 proved to be unsuccessful and instead led to degradation of the product
(entry 5). In hopes to prevent the formation of the acid labile orthoester the temperature of the
glycosylation was attempted at 40 oC (entry 6). Under these conditions the formation of the
orthoester was prevented, however, the disaccharide 18 was obtained in low yields. Surprisingly
in these conditions a new disaccharide 20 was also formed in low yields. This disaccharide was
determined to be the dechloroacetylated α-linked disaccharide 20. Proton NMR showed a
downfield shift of H-1 of the glucose residue to 5.12 ppm compared to 4.61 ppm in disaccharide
18. α-Linkages are known to shift the anomeric proton to a higher ppm compared to their β-
32
counterparts.24,48 Additionally, the JH-1,H-2 of this proton was 4.0 Hz, which is in the expected
range for a α-glycosylic linkage. The chloroacetyl group was determined to be cleaved by the
disappearance of its characteristic peaks on both the 1H and 13C NMR spectra, namely its proton
peaks around 3.3 ppm as well as the 2o carbon peak around 40.2 ppm and the 4o carbon peak
around 170 ppm. Furthermore, the H-2 signal on the glucose residue has shifted upfield to 3.68
ppm compared to 5.01 ppm in the β-linked disaccharide. This upfield shift is the result of the loss
of the withdrawing protecting group. The exact mechanism of how this reaction occurred is not
fully understood. One explanation is the migration of the chloroacetyl group. It has been reported
of acetyl groups migrating during glycosyations as well as in our group we have observed the
migration of acetyl groups to the anomeric carbon in acidic conditions.49,50 It is hypothesized that
in the more forceful conditions, the chloroacetyl group migrated to the anomeric position which
was activated by the BF3•OEt2 and glycosylated with the acceptor similarly to the synthesis of
the thiophenyl donor 4. The best conditions to synthesis the disaccharide showed to be catalytic
amounts of TMSOTf at room temperature which gave the disaccharide in moderate yields (entry
1).
33
Table 2. Glycosylation conditions for the glycosylation reaction of mannosyl acceptor 3 and
glycosyl donor 5.
Entry
Activator
(Equiv)
1
TMSOTf
(0.04)
TMSOTf
(0.04)
TMSOTf
(0.2)
BF3•OEt2
(0.2)
BF3•OEt2
(0.2 to 0.4)
BF3•OEt2
(0.2)
2
3
4
5
6
5
3
Equiv. Equiv.
Temp
4Å
Sieves
18
19
20
1
1.4
rt
No
54%
-
-
1.3
1
rt
-
85%
-
1.2
1
-78 to rt
18%
29%
-
1
1.3
rt
200
mg/mL
7
mg/mL
No
31%
-
-
1.5
1
rt
No
-
6%
-
1
1.3
40 oC
No
20%
-
10%
2.3.3 Synthesis of disaccharide acceptor 26.
Disaccharide 18 was dechloroacetylated in pyridine/ethanol with thiourea to give the
alcohol 21 in excellent yields. Inversion of O-2 of glucose was originally planned to be
accomplished by oxidation and reduction following the procedure described by Wu and Bundle,
which began with an overnight Swern-type oxidation with DMSO and acetic anhydride followed
by reduction using L-selectride®.38,51 However, in these conditions an unexpected side reaction
occurred resulting in 23 being the major product. This was identified by 1H and 13C NMR. The
absence of the H-3 on the glucose residue suggests an elimination occurred. Additionally, this is
also observed in the coupling pattern of H-4 as it is a doublet where it previously was a triplet.
Furthermore, H-4 has shifted downfield to 5.76 ppm due change in the carbon’s hybridization.
This downfield shift is also observed in the 13C NMR as C-3 appears as a 4o carbon at 147.0
ppm.
34
Scheme 23: Hypothesized mechanism for the formation of 24.
The formation of this alkene is believed to occur due to the basic conditions of the
reaction. The oxidation occurs as described, however, this increases the acidity of the
neighbouring proton. With excess AcO- present in solution the carbohydrate likely undergoes an
E2 type elimination with the deprotonation of H-3 with the acetyl protecting group leaving
resulting then α,β unsaturated ketone 23 (Scheme 23).
Additionally, difficulties with this reaction arose with the L-selectride®. L-selectride®
was shown to be too nucleophilic and would deprotect the acetyl groups of 21 at room
temperature. It was determined that lower concentrations of acetic anhydride (2 mM compared to
the 4 mM) and lowering the reaction time to five hours, allowed for the oxidation to work in
good yields. Additionally, degradation was minimized during reduction by keeping the reaction
at -78 oC giving the rhamnose disaccharide 22 in 45% over the two steps. The free hydroxyl
group was benzoylated with benzoyl chloride and pyridine to give the fully protected 24. The
axial hydroxyl was less reactive compared to other hydroxyl group requiring heat for protection
35
to occur. Methanoic HCl allowed for the selective deprotection of the acetate in the presence of a
benzoates to give the disaccharide acceptor 26 in good yields.
Scheme 24: Synthetic steps using to prepare disaccharide acceptor 26.
2.3.4 Synthesis of trisaccharide 27
The trisaccharide glycosylation with mannosyl donor 4 was conducted with NIS and
TfOH in poor yields. Due to lack of time this glycosylation was attempted only once on a small
scale. However, the results look promising and we expect the yields to increase upon
optimization.
36
Scheme 25: Glycosylation reaction to for trisaccharide 27.
2.4 Conclusion
Using the monosaccharide building blocks 3, 4 and 5 trisaccharide 27 has been
synthesized. Formation of the β-rhamnose linkage was achieved with neighbouring group
participation followed by inversion afterwards. It has been shown that ester protecting group are
not ideal for an Albright-Goldman oxidation. The basicity of the acetate anion was shown to be
able to deprotonate an α-hydrogen, which with the departure of the acetate group leads to an α,
β-unsaturated ketone forming. Additionally, ester protecting groups are labile in the presence of
borohydrides making the reduction quite difficult. Exploration into various ether protecting
groups might be advantageous to the synthesis of 27. Due to small quantities available, the
synthesis of the trisaccharide 27 was never optimized but it is hypothesized that this trisaccharide
can be synthesized in high yields.
37
Chapter 3: Synthesis of L-rhamnose trisaccharide fragment
38
3
Introduction
This chapter will focus on the synthesis of the L-rhamnose trisaccharide 2. The L-
trisaccharide proved to more difficult to synthesis than the D-counterpart. The primary difficulty
was the formation of the β-rhamnose glycosidic linkage. Unfortunately, the L-trisaccharide has
not been synthesized yet due to various setbacks in its synthesis. This chapter will discuss the
failed synthetic attempts to form trisaccharide 2 and the progress made towards the trisaccharide.
Scheme 26: Monosaccharide building blocks being used for the synthesis of 2.
3.1
Glycosylations with the 2, 3-O-carbonate rhamnosyl donor 32
Synthesis of trisaccharide 2 will involve mannosyl donor 4 and mannosyl acceptor 3
whose synthesis have already been previously described. Our initial synthetic plan was to use the
carbonate glycosyl halide 32 with insoluble silver catalysis. As previously mentioned these types
of donor are known to be β-directing. Synthesis began with commercially available L-rhamnose
monohydrate. Acetylation of L-rhamnose by pyridine and acetic anhydride in gave the known
39
peracetylated rhamnose in quantitative yields.52 The peracetylated rhamnose was glycosylated
with thiophenol and BF3•OEt2 to give the thioglycoside in 73% yield. Despite neighbouring
group participation 1H NMR analysis showed an anomeric mixture of 4:1 α:β was obtained.53
Deacetylation under Zemplèn conditions gave the triol 29 in quantitative yields. Formation of the
2,3-O-carbonate was accomplished with phosgene and pyridine. This allowed for the separation
of anomers giving exclusively the known α-anomer 30 in 61% yield. Despite following the
procedure described by Crich et al. our yields were anywhere from 20-30% lower than
described.54 This drop in yield is likely due to the presence of the β-glycoside either degrading or
undergoing a side reaction at this step. The O-3 alcohol was protected with a chloroacetyl group
which gave the novel fully protected thiorhamnoside in 88% yield. The chloroacetyl group was
selected as it was hoped to be selectively removed in the presence of carbonate group.
Conversion of the thioglycoside to the anomeric bromide 32 was accomplished with elemental
bromine and gave the donor in quantitative yields.
Scheme 27: Synthetic steps for the synthesis of bromide 32.
Many synthetic trials reactions were conducted with donor 32 to synthesis the βdisaccharide 33, which are summarized in table 3 but it was to no avail. Initial attempts of
40
glycosylation with insoluble silver catalysts, Ag2O and Ag2SiO3 at room temperature saw little to
no activation of the bromide (entries 1 & 2). When heated, activation was observed for Ag2SiO3
but not Ag2O. However, the disaccharide 34 formed was via a α-linkage (entry 3). This was
confirmed by the rhamnose residue’s 1JC-1, H-1, value, which was 169.5 Hz and is indicative of a
α-linkage. Additionally, H-1 of the rhamnose residue appeared as singlet which coincides with
what Crich et al. reported where α-anomers 3JH-1, H-2 was 0 Hz in the α-anomer and
approximately 3.0 Hz for the β-anomer.34
Scheme 28: Glycosylation reaction of mannosyl acceptor 3 and rhamnosyl bromide 32.
The formation of this disaccharide is hypothesized to be formed as a result of the
degradation of the glycosyl halide. This degradation creates HBr, which in situ activated 32 and
formed the α-disaccharide in small quantities. Betaneli et al. have reported similar difficulties in
forming the β-linkage with silver oxide.55 Their solution was the addition of catalytic amounts of
41
the silver perchlorate to the reaction which proceeded to the β-linkage as the favoured product.
Due to the safety concerns with AgClO4 we instead chose to use AgOTf as a soluble silver salt
but unlike Betaneli et al. only an α-linked was observed (entry 4). It is likely that the small
quantities of soluble silver was able to activate the bromide donor, removing β-selectivity.
Table 3. Glycosylation reaction conditions of mannosyl acceptor 3 and rhamnosyl bromide 32.
Entry
Activator
Solvent
Temp.
34
1
Ag2O
DCM
rt or 40 oC
NR
2
Ag2SiO3
DCM
rt
NR
3
Ag2SiO3
DCE
50 oC
6%
4
Ag2SiO3, AgOTf
DCM
rt
25%
5
Ag2SiO3, AgOTf
MeCN
rt
17%
6
Hg(CN)2
MeCN
50 oC
15%
After our failed attempts in halogenated solvents the glycosylation was performed in a βdirecting solvent. Acetonitrile has been shown to be a β-directing solvent as it coordinates axially
to the oxocarbenium ion forming the nitrilium ion resulting in the β-glycoside being favoured
(Scheme 29).56 We conducted the previous AgOTf glycosylation conditions (entry 4) again but
with acetonitrile as the solvent. Additionally, Hg(CN)2 was tried as an activator in acetonitrile,
however, both glycosylations gave the α-disaccharide in low yields (entry 5 & 6). Overall,
despite our best efforts formation of the desired product was never obtained. This is likely due to
the low reactivity of the acceptor. As reported by Paulsen, activations with insoluble silver
42
catalysts are limited in use.29 Typically, insoluble silver catalysis works for primary alcohols or
highly reactive secondary nucleophiles, and the effectiveness decreases as nucleophilicity of the
acceptor decreases.
Scheme 29: The β-directing effect of the nitrilium ion.13
3.2
Glycosylation with neighbouring group participation
3.2.1 Inversion attempts with benzylated rhamnosyl triflate 39
Since we were unable to form a β-linkage with the carbonate donor alternate strategies
were attempted to synthesize the disaccharide. It was decided that neighbouring group
participation would be used to ensure a β-glycosidic linkage which would be inverted back to Lrhamnose afterwards. However, with L-glucose being quite rare and expensive it would be
synthesized from L-rhamnose via an SN2 substitution. The synthesis began with commercially
available L-rhamnose and through a Fischer glycosylation the anomeric oxygen was
functionalized to an allyl glycoside (Scheme 30). The 2,3-cis diol was protected with an
isopropylidene ring using acetone and DMP to give the known alcohol 35 in good yields.57
Protection of O-4 was accomplished with a levulenic ester to giving 36. The levulenic ester was
chosen as it can be selectively deprotected in the presence of other esters with hydrazine acetate.
Hydrolysis of the isopropylidene occurred in 80% acetic acid at 80 oC giving diol 37. Formation
of the dibutyltin acetal was accomplished in toluene, and upon treating with CsF and benzyl
43
bromide gave exclusive formation of the O-3 ether 38. Triflation of O-2 with triflic anhydride
and pyridine gave the protected triflate 39 in excellent yields.
Scheme 30: Synthetic steps to rhamnosyl triflate 39.
The attempts to invert 39 are summarized in Table 4. Initially, the acetate anion was used
as the nucleophile, however, instead of a substitution an elimination occurred between C-2 and
C-3 giving 41. The elimination occurred at both 60 oC and room temperature. The eliminated
product was confirmed through 1H and 13C NMR analysis. In the l3C NMR spectrum C-3 had a
large upfield shift which was observed at 154 ppm and as a quaternary carbon compared to C-3
of the starting material which is at 74 ppm. C-2 of 41 also had a noticeable shift downfield to 95
ppm from 71 ppm. Furthermore, there were distinct differences in the proton NMR. The
eliminated sugars had one fewer proton than the triflate. Additionally there are significant
changes to the coupling patterns in the eliminated product. Both H-1 and H-2 show as doublets
with a 2.7 Hz coupling constant compared to the broad singlets found in the precursor. The
change in the coupling pattern is likely due to the double bond distorting the ring. This distortion
44
resulted in alteration of the dihedral angle and altering the coupling constant.58 The coupling
pattern of H-4 differed from the triflate 39. H-4 of 39 was a triplet with a 9.9 Hz coupling
constant, while H-4 in 40 was a doublet with a coupling constant of 6.9 Hz. The change in
coupling pattern is due to the loss of the H-3 proton and causing the H-4 to only split once, and
the change in the coupling constant is likely due to the previously mentioned ring distortion.
We determined that the acetate anion was too basic and not nucleophilic enough for this
reaction to work as desired. To avoid the elimination we attempted a Lattrell-Dax nitrite
mediated inversion.59 The Lattrell-Dax inversion was attempted to invert the triflate to a
hydroxyl group but instead a complex mixture formed with the desired product 40 being attained
in small yields. These results coincided with the work conducted by Dong et al.60 They showed
that neighbouring benzyl groups in a Lattrell-Dax inversion resulted in a complex mixture.60 The
complex mixture is a result of the donating nature of the benzyl group. This makes the
neighbouring hydrogen too reactive and leads to multiple side reactions such as ring contractions
and eliminations. They noted that if a neighbouring ester group was present that the inversion
occurred successfully.
45
Table 4. Synthetic attempts to invert rhamnosyl triflate 39.
3.2.2
Entry
Reagent
Temp
Equiv.
Yield of 40
Yield of 41
1
KOAc
40 oC
8
-
88%
2
TBAOAc
rt
8
-
85%
3
KNO2
50 oC
8
-
10%
4
KNO2
rt
8
-
-
5
TBANO2
50 oC
8
4%
12%
6
TBANO2
rt
8
-
11%
7
TBANO2
rt
30
3%
10%
Inversion of benzoyl rhamnosyl triflate 43
From this we synthesized the benzoate analogue 43 (Scheme 31). Initially, formation of
42 from the diol 37 was attempted through a stannylene acetal, however, this gave a mixture of
monobenzoate products with the O-2 benzoate as the major product. Selective benzoylation of
O-3 was achieved in good yields using limited equivalents of BzCl in pyridine. Triflation of the
alcohol gave the triflate 43 in excellent yields.
46
Scheme 31: Synthetic steps to several rhamnose sulfonates 43, 44 and 45.
The result of the inversion attempts are summarized in Table 5. Attempts to invert with
tetrabutylammonium acetate resulted in an elimination similar to 39 (entry 1). The Lattrell-Dax
nitrite mediated inversion was then attempted in multiple conditions. Whether the inversion was
attempted with 5 equivalents of nucleophile at elevated temperatures or in large excess of
nucleophile at room temperature the outcome of the reaction was the same. Both conditions gave
the desired product 46 and the eliminated product 47 in low yields in almost a 1:1 ratio (entries 3
& 4). The eliminated product was identified similarity to the previously mentioned 41. The
inverted sugar was determined by the change in coupling constants/coupling patterns. Inversion
at C-2 changes the dihedral angle between H-1, H-2 and H-2, H-3, which can be seen in the
coupling pattern of H-2. After inversion H-2 shows as a doublet of doublets with JH-1, H-2 = 3.2
Hz and JH-2, H-3 = 9.6 Hz where previously it was a broad singlet. Additionally, H-2 has an upfield
shift to 3.79 ppm from 5.17 ppm as a result of the loss of the triflate. Due to the low basicity of
the nitrite ion it is likely that the elimination occurs through an E1 mechanism. We then
hypothesized that by reducing the reactivity of the leaving group then the overall chances of
elimination would be reduced. Therefore the triflate was replaced with a mesylate group and
tosylate group (Scheme 31). However, both sulfonyl groups failed to invert under the previous
conditions. The inversion was attempted with heat increasing upwards to 100 oC, but the sugars
47
degraded before any inversion was observed (entries 6 & 7). We determined that these
alternative leaving groups were not reactive enough for this purpose.
Table 5. Synthetic attempts to invert rhamnosyl sulfonates 43, 44 and 45.
Entry
X
(R)
Temp
Equiv.
Yield of 46 Yield of 47
1
OTf
AcO-
rt
5
-
77%
2
OTf
NO2-
rt
5
-
-
3
OTf
NO2-
50 oC
5
23%
27%
4
OTf
NO2-
rt
25
23%
28%
5
OTf
NO2-
rt →50 oC
25
25%
30%
6
OMs
NO2-
rt to 110 oC
25
-
-
7
OTs
NO2-
rt to 110 oC
25
-
-
3.3 Glycosylations with non-participating sulfonyl groups.
It has been reported that non-participating sulfonyl groups have been shown to be
potentially β-directing.61 It was hypothesized that the triflate would be too reactive and would
degrade during the hydrolysis of the aglycone or during glycosylation. Instead two donors with
either the O-2 mesylate 50, or O-2 tosylate 51, were synthesized (Scheme 32). Removal of the
allyl groups occurred in two steps, initially by converting the aglycone to a vinyl glycoside,
which was hydrolyzed to the hemiacetal using aqueous mercury salts. The hemiacetal was
48
converted to the trichloroacetimidate donors 50, 51 with trichloroacetonitrile and DBU in good
yields.
Scheme 32: Synthetic steps to rhamnosyl trichloroacetimidates 50 and 51.
Glycosylation was attempted with the mannosyl acceptor 3 with both donors (Scheme
33). In both cases formation of the disaccharide was achieved, however, both gave αdisaccharides 51 and 52. These were determined by the J1C-H which was 174 Hz in both cases.
With the formation of the α-linkage we determined that these non-participating sulfonyl groups
have minimal β-directivity.
Scheme 33: Glycosylation reaction of mannosyl acceptor 3 and rhamnosyl sulfonate 44 and 45.
49
3.4 Conclusion
We experienced much difficulties in the synthesis of a β-mannoside, and despite our best
efforts this was not accomplished. We have attempted several synthetic pathways to the βmannoside with little to no success. The 2,3-O-carbonate protecting group was shown to lower
the reactivity of the glycosyl bromide. However, in the presence of an insoluble catalyst and poor
nucleophile bromide 32 did not react. The use of more forceful conditions or soluble silver
catalysts did result in a glycosylation occurring but with no β-selectivity. While O-2 nonparticipating electron disarming protecting groups have shown to be potentially β-directing, this
was not observed in this research. Both the O-2 mesylate and O-2 tosylate were shown not to be
β-directing. Inversion at O-2 of rhamnose also proved to be quite difficult. Less labile sulfonyl
leaving groups are not reactive enough to undergo an inversion, while the triflate was too
reactive and resulted in a mixture of products forming with the desired product only forming in
low yields. However, inversion with triflate 43 has shown to be the most promising synthetic
pathway. Currently, we plan to synthesize the β-mannoside through an inversion at O-2. Despite
the poor yields, the inversion is the best way to ensure the β-linkage. The work is currently
underway in our lab and the proposed synthesis to the trisaccharide will be discussed in the next
chapter.
50
Chapter 4: Future work
51
4.1 Synthesis of L-trisaccharide 59
Currently, there is several projects underway within our group. To increase our overall
quantity of synthesis of the D-rhamnose trisaccharide 27 is underway. Additionally, synthesis of
the L-analogue stills needs to be accomplished with current plans being to go through the
inversion despite the poor yields. Our synthetic strategy begins with the acetylation of 46 to
allow for participation during glycosylation (Scheme 34). Deprotection of the anomeric allyl
group will follow the previous procedures and the trichloroacetimidate will be introduced.
Glycosylation with the mannose acceptor will hopefully yield disaccharide 56. Removal of the
levulenic ester will occur with hydrazine acetate giving disaccharide acceptor 57 which will be
glycosylated with the mannosyl donor 4 giving trisaccharide 58. The synthesis will be completed
with inversion of O-2 through an oxidation/reduction giving the protected CPS fragment 59.
Scheme 34: Proposed synthetic steps to trisaccharide 59.
52
4.2 Deprotection and NOE analysis
With both fully protected trisaccharides global deprotection in dissolving metal
conditions will give the target trisaccharides 1 and 2. Once we have the trisaccharides, NMR
experiments will be conducted and compared to the reported spectra. Pequegnat et al. have
already conducted 1H and 1H NOESY experiments to which we will compare our results with
their findings to confirm the absolute configuration of the rhamnose residue.9
Scheme 35: Global deprotection of trisaccharides 28 and 59 giving CPS fragments 1 and 2.
The NOE experiments previously ran by Pequegnat et al. showed interesting interactions
for H-6 of the rhamnose unit (Figure 13). Crosspeaks were observed between H-6 Rha-H-1 Man
as well as H-6 Rha-H-2 Man and H-6 Rha-H-3 Man. It is believed that these specific interactions
would depend highly on the absolute configuration of the rhamnose residue. Therefore, it is
53
believed that NOE spectra will be unique to the specific enantiomer and that comparison to the
previous results will confirm the absolute configuration of the rhamnose residue.
Figure 13: NOE interactions of Clostridium bolteae CPS. Taken from Pequegnat et al.9
4.3 Disaccharide fragments 60 and 61
Since the interactions of interest is across the nearby 1-4 α linkage, therefore, formation
of the difficult β-linkage might not be necessary. We hypothesize that the conformation of a
disaccharide fragment would not differ significantly to the trisaccharide. Due to this the synthesis
of two disaccharides with different rhamnose configurations may be able to confirm the absolute
configuration of the rhamnose unit. Currently within our group the synthesis of 60 and 61 is
already underway. These disaccharides will hopefully provide more information of the structure
of the Clostridium bolteae CPS.
54
Figure 14: Target disaccharide Clostridium bolteae CPS fragments 60 and 61.
4.3.1 Synthetic strategy for the synthesis of 60
Synthesis of the D-analogue will follow the same synthetic pathway as the quinovosyl
donor until 15 (Scheme 36). Protection of the alcohol will be changed to an allyl group from an
acetyl group. We decided to change to the allyl protecting group in hopes to improve the yield at
the inversion step. The allyl group is a much poorer leaving group than the acetate, therefore,
occurrence of the elimination during the oxidation is less likely. Additionally, the allyl group is
resistant to base preventing degradation during the reduction step. From here, the synthesis
follows a similar pathway as the quinovosyl donor where it will be equipped with a methyl
glycoside via a β-linkage. The sugar will then undergo a Zèmplen deacetylation, which will
undergo an oxidation, followed by reduction to give the rhamnose unit 66. Deprotection of the
allyl group will give acceptor 67 which will be glycosylated giving the protected disaccharide
60.
4.3.2 Synthetic strategy for the synthesis of 61
Regarding the L-disaccharide, synthesis will begin with thiophenyl rhamnose and will be
protected with an isopropylidene group giving us the known rhamnosyl acceptor 68.
Glycosylation with perbenzoylated mannosyl bromide will give disaccharide 69. Silver triflate
will be the activator in this glycosylation to avoid the formation of the orthoester. Hydrolysis of
55
the isopropylidene will give the diol, which will be converted to the carbonate 70 using
phosgene. Conversion of the thiol to a bromide using bromine to which a glycosylation will be
attempted using insoluble silver catalysis (Scheme 37). It is believed that the higher
nucleophilicity of methanol compared to 3 will lead to the formation of the β-glycoside.
Scheme 36: Proposed synthetic steps to CPS fragment 60.
Global deprotection of disaccharides 60 and 61 will occur similarly to the trisaccharides 1
and 2 comparison to the NOE will occur as discussed above. With this additional information the
absolute configuration of the rhamnose residue can be determined. Confirmation of the absolute
configuration of the rhamnose unit allows for the synthesis of larger CPS fragments to begin.
With larger CPS fragments immunological screening of a potential anti-C. bolteae vaccine can
occur.
56
Scheme 37: Proposed synthetic steps to CPS fragment 61.
57
Chapter 5-Experimental
58
1
H NMR (400 or 600 MHz, 295 K) and 13C NMR (100 MHz or 150 MHz, 295 K) spectra were
recorded in CDCl3 (calibrated at δC 77.0 ppm, and using residual CHCl3 at δH 7.24 ppm). All
chemical shifts are reported in parts per million (ppm). All coupling constants (J) are reported in
hertz (Hz) and were obtained from analysis of 1H NMR spectra or uncoupled HSQC spectra.
Assignments of proton and carbon peaks were made by analysis of 2D COSY and HSQC
experiments. Multiplicities are abbreviated as singlet (s), broad singlet (bs), doublet (d), doublet
of a doublet (dd), doublet of a doublet of a doublet (ddd), triplet (t), doublet of a triplet (dt),
triplet of a doublet (td), triplet of a triplet (tt), quartet (q), doublet of a quartet (dq), quartet of a
doublet (qd), quintet (quin), and multiplet (m). Anhydrous solvents were freshly distilled.
Organic solutions were dried over Na2SO4 and concentrated under reduced pressure. All
products were dried over high vacuum. Molecular sieves were activated by heating at 300 °C
under reduced pressure. Thin layer chromatography (TLC) was performed on aluminum plates
coated with silica gel and charred with 20% H2SO4 in EtOH. Compounds were purified by
column chromatography using silica gel unless otherwise stated. Reverse phase HPLC
purifications were carried out on a Prep Nova Pak HR C18, 6 μm 60 Å column using HPLC
grade acetonitrile and milli-Q water. Optical rotations were measured at 22 °C on a Rudolph
Research Autopol III polarimeter and reported as follows: [α]D (c in g per 100 mL, solvent).
High resolution electrospray ionization mass spectra (HRESIMS) or high resolution electron
ionization (HREI-MS) were recorded by the analytical services of the Queen's Mass
Spectrometry and Proteomics Unit, Kingston, Ontario.
59
3-O-Benzyl-1,2-O-isopropylidene-α-D-glucofuranose (10)
Diacetone-D-glucose (12.36 g, 47.5 mmol) was dissolved in anhydrous THF (100 mL) with
BnBr (11.00 mL, 1.95 equiv) and TBAI (0.16 g, 0.01 equiv) under N2 and cooled to 0 oC. NaH
(60% dispersion 6.85 g, 3.6 equiv) was added slowly over 5 min and stirred overnight at room
temperature. The reaction was then diluted with EtOAc (100 mL) was washed with water (2 ×
100 mL). The solution was dried over Na2SO4, filtered and concentrated to an yellow oil. The
solution was dissolved in 80% and stirred overnight at room temperature. The solution was coconcentrated with toluene (3 × 100 mL) and the product was purified by flash chromatography
(1:1 EtOAc:Hexanes) to give the known diol 10 (12.82 g, 87%) over the two steps as a yellow
oil. 1H NMR for 10 (CDCl3, 400 MHz, 295 K) δH 7.35-7.30 (m, 5H, Bn), 5.59 (d, 1H, J = 3.9
Hz, H-1), 4.73 (d, 1H, J = 12.6 Hz, CHHAr), 4.62 (d, 1H, J = 3.9 Hz, H-2), 4.53 (1H, J = 12.6
Hz, CHHAr), 4.11-4.07 (m, 2H, H-3, H-4), 4.02 (m, 1H, H-5), 3.80 (d, 1H, J = 11.5 Hz, H-6a),
3.68 (dd, 1 H, J = 5.5 Hz, 11.5 Hz, H-6b), 2.53 (bs, 1H, OH-5), 1.46 (s, 3H, OCCH3), 1.30 (s,
3H, OCCH3). The NMR data are in agreement with those reported in the literature.42
60
3-O-Benzyl-1,2-O-isopropylidene-6-O-tosyl-α-D-glucofuranose (11)
Diol 10 (9.37 g, 30.2 mmol) was dissolved in anhydrous DCM (350 mL) and pyridine (10 mL,
4.27 equiv) and cooled to -20 oC. The solution was left stirring for 1 hr and then TsCl (5.5 g 0.99
equiv) was added and allowed to warm to RT. After 3 h the solution was cooled back down to 20 oC and an addition of TsCl (1.65 g, 0.3 equiv) and DMAP (350 mg, 0.1 equiv) was added.
The solution was kept cold for an additional 1 h and then allowed to warm to RT and stirred
overnight. The solution was quenched with 2 M HCl (100 mL) then extracted and washed with
saturated NaHCO3 (100 mL). The solution was dried over Na2SO4, filtered and concentrated to
an yellow oil. The product was purified by flash chromatography (1:1 EtOAc:Hexanes) to give
the known tosylate 11 (11.78 g. 85% yield) as a clear oil. 1H NMR for 11 (CDCl3, 400 MHz,
295 K) δH 7.76 (d, J = 8.5 Hz, SO2Ar) 7.36–7.26 (m, 7 H, Bn, SO2Ar), 5.86 (d, 1 H, J = 4.0 Hz,
H-1), 4.68 (d, 1 H, J = 11.7 Hz, CHHAr), 4.57 (d, 1H, J = 3.7, H-2) 4.54 (d, 1 H, J = 11.7 Hz,
CHHAr) 4.24 (dd, 1 H, J = 2.8 Hz ,10 Hz, H-6a), 4.17 (m, 1 H, H-5), 4.13 (t, 1 H, J = 3.3 Hz, H3), 4.05-4.01 (m, 2 H, H-4, H-6b), 2.60 (d, 1 H, J = 5.8 Hz, OH-5), 2.41 (s, 3 H, CH3C6H4),
1.43 (s, 3H, OCCH3), 1.28 (s, 3H, OCCH3). The NMR data are in agreement with those reported
in the literature.43
61
6-Anhydro-3-O-benzyl-α,β-D-glucopyranose (12)
Ketal 11 (11.50 g, 24.05 mmol) was dissolved in dry THF (150 mL) and LiAlH4 (3.65 g, 4
equiv) was added slowly over 5 minutes and the reaction was stirred for an additional 15 min.
The reaction was then quenched by dropwise addition of ice cold H2O (5 mL). The solution was
further diluted with 15% NaOH (5 mL) and water (5 mL) and stirred for 30 min. The solution
was filtered over celite and washed with EtOAc (3 × 25 mL). The filtrate was separated and
dried over Na2SO4 and concentrated to grey oil. The crude sugar was dissolved in 80% AcOH
(200 mL) and was heated to 80 oC overnight. The solution was co-concentrated with toluene (3 ×
150 mL) and the product was purified by flash chromatography (8:2 EtOAc:Hexanes) to give the
known triol 12 (4.32 g, 71% over the two steps, 1:1 α:β) as a clear oil. 1H NMR for 12 (CDCl3,
400 MHz, 295 K) 7.40-7.25 (10 H, Bn α/β) 5.20 (1H, t, J = 3.0 Hz, H-1α), 4.99 (d, 2H, J = 11.5
Hz, CH2Bn α/β), 4.75 (dd, 2H, J = 11.5 Hz, CH2Ar α/β), 4.58 (1H, dd, J = 5.5, 7.1 Hz, H-1β),
3.92-3.88 (m, 1H, H-5β), 3.69 (td, 1H, J = 3.1, 9.7, 12.3 Hz, H-2α), 3.59 (t, 1H, J = 9.3 Hz, H3β), 3.48 (td, 1H, J = 1.7, 9.3 Hz, H-2β), 3.41-3.37 (m, 1H, H-5α), 3.36 (t, 1H, J = 8.4 Hz, H-3α),
3.27 (td, 1H, J = 2.6, 8.8 Hz, H-4β), 3.21 (td, 1H, J = 3.1, 8.8 Hz, H-4α), 3.15 (d, 1H, J =5.8 Hz,
OH-1β), 2.82 (d, 1H, J =2.8 Hz, OH-1α), 2.39 (d, 1H, J =1.8 Hz, OH-2β), 2.19-2.12 (m, 3H,
OH-2α, OH-4β, OH-4α), 1.30 (d, 3H, J = 5.8 Hz, H-6β), 1.25 (d, 3H, J = 6.6 Hz, H-6α). The
NMR data are in agreement with those reported in the literature.44
62
6-Anhydro-3-O-benzyl-1,2-di-O-chloroacetyl-α,β-D-glucopyranose (13)
Triol 12 (630 mg, 2.5 mmol) was dissolved in dry DCM (20 mL) and Et3N (3 mL, 20.7 mmol)
and cooled to -20 oC under nitrogen. ClAcCl was then added dropwise (430 μL, 2 equiv) and
stirred for 1h. An addition of ClAcCl (11 μL, 0.5 equiv) was added every hour until 3.5 equiv of
ClAcCl was reached. The solution was quenched with 5 mL H2O was washed with 25 mL 1N
HCl followed by 25 mL of sat’d NaHCO3. The organic layers were dried over Na2SO4 and
concentrated to dichloroacetyl 13 (824 mg, 52%, 3:1 β:α) a yellow oil. 1H NMR for 13 (CDCl3,
400 MHz, 295 K) δH 7.37-7.24 (m, 10H, Bn α/β), 6.29 (d, 1H, J = 3.6 Hz, H-1α), 5.66 (d, 1H, J
= 8.3 Hz, H-1β), 5.11 (dd, 1H, J = 8.3, 9.0 Hz, H-2β), 5.04 (dd, 1H, 3.6 Hz, 10.0 Hz, H-2α), 4.81
(d, 1H, 12.1 Hz, CH2Bn α) 4.73 (m, 3H, CH2Ar α/β), 4.10 (s, 2H, ClAc α), 4.04 (d, 2H, 1.4 Hz,
ClAc β), 3.93-3.76 (m, 6H, ClAc α/β, H-3α, H-4α), 3.57 (m, 2H, H-3β, H-5β), 3.38-3.31 (m, 2H,
H-4α/β), 2.29 (d, 1H, J = 3.0 Hz, OH-4α), 2.18 (d, 1H, J = 3.4 Hz, OH-4β), 1.32 (d, 3H, J = 6.1
Hz, H-6β), 1.29 (d, 3H, 6.2 Hz, H-6α). 13C NMR (CDCl3, 100 MHz, 295 K): δC 166.0, 165.9 (2
× C=O α,β), 137.7, 137.5 (4o Ar α,β), 128.8-127.7 (Ar α,β), 92.9 (C-1β), 91.4 (C-1α), 82.1 (C3α), 78.8 (C-3α), 75.3 (CH2Ar α), 75.1 (CH2Ar β), 74.9 (C-4β), 74.6 (C-4α), 73.7 (C-2α), 76.6
(C-2β), 72.9 (C-5 β), 72.7 (C-5α), 40.5-40.2 (2 × CH2Cl α,β) 17.4 (C-6 α,β). HRESIMS
calculated for C17H20Cl2O7 [M+Na]+ 429.0484, found 429.0487.
63
4-O-Acetyl-6-anhydro-3-O-benzyl-1,2-O-isopropylidene-α-D-glucopyranose (16)
Triol 12 (4.75 g, 18.7 mmol) was dissolved in anhydrous THF (70 mL) and DMP (9.25 mL, 75.5
mmol) and stirred for 15 minutes. CSA (894.5 mg, 3.85 mmol) was added and the reaction was
stirred under nitrogen for 40 hours. The reaction was quenched with Et3N (6.85 mL) and
concentrated to an yellow oil. The oil was dissolved in a 1:1 mixture of Ac2O:Pyr (120 mL) and
stirred for 90 minutes at room temperature. The reaction was co-concentrated with toluene (3 ×
75 mL) and was purified by flash chromatography (20% EtOAc:Hexanes) to give the acetylated
16 (5.14 g, 81% over the two steps) as an oil. [α]D 45.6 (c 1.0, MeOH) 1H NMR for 16 (CDCl3,
400 MHz, 295 K) δH 7.32-7.24 (m, 5H, Bn), 5.55 (d, 1H, J = 5.2 Hz, H-1), 4.77-4.74 (m, 2H, H4, CHHAr), 4.66 (AB pattern, 1H, J = 12.4 Hz, CHHAr), 4.19 (t, 1H, J = 4.4 Hz, H-2), 3.96-3.84
(m, 1H, H-5), 3.68 (t, 1H, J = 3.2 Hz, H-3), 2.05 (s, 3H, OAc), 1.53 (s, 3H, OCCH3), 1.32 (s, 3H,
OCCH3), 1.22 (d, 3H, J = 6.4 Hz, H-6) 13C NMR (CDCl3, 100 MHz, 295 K): δC 170.2 (C=O),
137.7 (4o arno), 129.0 (Bn), 128.8 (Bn), 125.6 (Bn), 109.0 (OCCH3), 96.9 (C-1), 76.5 (C-3), 74.7
(C-4), 73.9 (C-2), 71 (CH2Bn), 64.9 (C-5), 26.5 (OCCH3), 25.5 (OCCH3), 20.9 (OC(O)CH3),
18.2 (C-6). HRESIMS calculated for C18H24O6 [M+Na]+ 359.1470, found 359.1450.
64
4-O-Acetyl-6-anhydro-3-O-benzyl-α,β-D-glucopyranose (17)
Acetal 16 (5.14 g, 15.2 mmol) was dissolved in 80% AcOH (240 mL) was stirred for 2.5 h at 80
o
C. The reaction was removed from heat and cooled to RT and then was co-concentrated with
toluene (4 × 125 mL). The crude was purified with silica chromatography (7:3 EtOAc:Hex) to
give the hemiacetal 17 (3.54 g, 78% 1:1.1 α/β) as a clear yellow oil. 1H NMR for 17 (CDCl3, 400
MHz, 295 K) δH 7.34-7.24 (m, 10H, Bn), 5.22 (d, 1H, J = 2.4 Hz, H-1α), 4.83-4.66 (m, 6H,
CH2Ar α/β, H-4 α/β), 4.56 (d, 2H, J = 6.4 Hz, H-1β), 4.02 (m, 1H, H-5), 3.75 (m, 2H, H-2α, H3α), 3.50 (m, 3H, H-2β, H-3β, H-5β), 1.98 (s, 3H, OCCH3 α), 1.97 (s, 3H, OCCH3 β) 1.18 (d, 3H,
J = 6.4 Hz, H-6β), 1.13 (d, 3H, J = 6.4 Hz, H-6α) 13C NMR (CDCl3, 100 MHz, 295 K): δC
170.0, 169.9 (C=O α, β), 138.1, 138.0 (4o Ar α, β), 128.4-127.6 (Ar α, β), 96.4 (C-1β), 92.1 (C1α), 81.3 (C-3β), 79.5 (C-3α), 74.8 (C-4α), 74.5 (CH2Ar α), 74.4 (C-4β), 74.3 (CH2Ar β), 72.4
(C-2α), 70.2 (C-2β), 65.9 (C-5β), 65.8 (C-5α), 20.9 (COCH3 α), 20.8 (COCH3 β), 17.3 (C-6β),
17.2 (C-6α). HRESIMS calculated for C15H20O6 [M+Na]+ 319.1157, found 319.1138.
65
4-O-Acetyl-6-anhydro-3-O-benzyl-1,2-di-O-chloroacetyl-α-D-glucopyranose (14)
Method A:
Alcohol 13 (1.26 g, 2.80 mmol) was dissolved in a 1:1 mixture of pyridine:acetic anhydride (100
mL). The reaction was stirred at RT for 15 and then was co-concentrated with toluene (4 × 75
mL). The crude was purified by silica chromatography (2:8 EtOAc:Hex) to give the acetylated
14 (894.6 mg, 71%) as an oil.
Method B:
Diol 17 (3.54 g, 11.9 mmol) was dissolved in anhydrous DCM (270 mL) and anhydrous pyridine
(5.80 mL, 6 equiv). The reaction was stirred at 0 oC for 30 min and then the ClAcCl (3.0 mL, 3.1
equiv) was added dropwise and the reaction was allowed to warm to RT. After 1hr of stirring
another addition of ClAcCl (1.0 mL, 1 equiv) occurred and stirred for another hour. The reaction
was quenched with H2O (50 mL). The water was extracted and the organic phase was washed
with 2 M HCl (100 mL) followed by sat’d NaHCO3 (50 mL). The organic phase was dried and
concentrated. The crude was purified by silica chromatography (2:8 EtOAc:Hex) to give the
acetylated 14 (4.86 mg, 91%, 1:1.1 β:α) as an oil. 1H NMR for 14 (CDCl3, 400 MHz, 295 K) δH
7.30-7.20 (m, 10H, Bn α,β), 6.33 (d, 1H, J = 3.4 Hz, H-1α), 5.66 (d, 1H, J = 8.4 Hz, H-1β), 5.16
(dd, 1H, J = 8.2, 9.4 Hz, H-2β), 5.10 (dd, 1H, J = 3.7, 9.9 Hz, H-2α) 4.93-4.87 (m, 2H, H-4α,β),
4.68-4.52 (m, 4H, CH2Ar α,β), 4.11 (bs, 2H, CH2Cl), 4.05 (bs, 2H, CH2Cl), 3.95-3.74 (m, 6H, 2
× CH2Cl, H-3α,β, H-5α,β), 2.01 (s, 3H, OC(O)CH3), 1.98 (s, 3H, OC(O)CH3), 1.22 (d, 3H, J =
6.1 Hz, H-6α), 1.18 (d, 3H, J = 6.3 Hz, H-6β). 13C NMR (CDCl3, 100 MHz, 295 K): δC 169.5,
66
169.1, 165.8, 166.1 (2 × C=O α,β), 137.9, 137.5 (4oArmo α,β), 128.6-127.7 (CArmo α,β), 92.8 (C1β), 91.0 (C-1β), 79.6 (C-3β), 76.4 (C-3α), 74.9 (CH2Ar α), 74.4 (CH2Ar β), 73.9 (C-4β), 73.6
(C-4α), 73.3 (C-2α), 73.2 (C-2β), 71.5 (C-5 β), 68.9 (C-5α), 40.5-40.2 (2 × CH2Cl α,β), 20.8
(C(O)CH3 α), 20.5 (C(O)CH3 β) 17.3 (C-6 β), 17.2 (C-6 α). HRESIMS calculated for
C19H22O8NCl2 [M+Na]+ 471.0594, found 471.0598.
.
4-O-Acetyl-6-anhydro-3-O-benzyl-2-O-chloroacetyl-1-O-trichloroacetimidate-α-Dglucopyranose (5)
Chloroacetyl 14 (5.01 g, 11.13 mmol) was dissolved in THF (65 mL) and DMAPA (1.67, 1.2
equiv) was added dropwise and stirred at room temperature. After 1 hr the starting material had
disappeared and the reaction was diluted with DCM (50 mL) and washed with 1 M HCL (20
mL) followed by a sat’d NaHCO3 wash (25 mL). The reaction was dried over sodium sulfate,
filtered and concentrated to a yellow oil (3.89 g crude). The crude was dissolved in anhydr.
DCM (90 mL) placed under a N2 balloon. Trichloroacetonitrile (4.5 mL, 3.5 equiv) and DBU
(500 μL, 0.25 equiv) were added sequentially and the reaction was stirred overnight at room
temperature. The reaction changed to a black solution overnight, which was concentrated and
purified by silicia chromatography (2:8 EtOAc:Hexane + 0.1% Et3N) to give the donor 5 (3.94 g,
68%) as a yellow oil. 1H NMR for 5 (CDCl3, 400 MHz, 295 K) δH 8.57 (s, 1H, NH), 7.32-7.26
(m, 5H, Bn), 6.45 (d, 1H, J = 3.6 Hz, H-1), 5.08 (dd, 1H, 3.6 Hz, 10.0 Hz, H-2) 4.92 (t, 1H, J
=9.8 Hz, H-4), 4.65 (s, 2H, CH2Ar), 4.04 (t, 1H, J = 9.6 Hz, H-3), 4.00-3.96 (m, 1H, H-5), 3.83
67
(AB pattern, 2H, 15.0 Hz, CH2Cl), 1.99 (s, 3H, OC(O)CH3), 1.18 (d, 3H, J = 6.3 Hz, H-6). 13C
NMR (CDCl3, 100 MHz, 295 K): δC 169.5 (C=O), 166.3 (C=O), 160.8 (C=NH), 137.8 (Bn),
128.4 (Bn), 127.9 (Bn), 127.8 (Bn), 93.1 (C-1), 76.3 (C-3), 74.9 (Bn), 74.1 (C-4), 73.9 (C-2),
68.7 (C-5), 40.2 (CH2Cl), 20.9 (OC(O)CH3), 17.3 (C-6).
Methyl 4,6-di-O-benzyl-2,3-O-isopropylidene-α-D-mannopyranoside (7)
Methyl α-D-mannopyranoside (5.00 g, 25.7 mmol) was suspended in a 1:1 ratio of acetone and
2,2-dimethoxypropane (100 mL) with p-TsOH (1.01 g, 0.2 equiv). The solution was stirred at 40
o
C for 3 h until all the sugar had dissolved. The solution was cooled to RT and then diluted with
water (100 mL) then stirred for 3.5 h. The solution was quenched with solid NaHCO3 (500 mg)
and the organic solvents were removed under vacuum. The aqueous layer was extracted by
hexanes (2 × 50 mL) and then chloroform (4 × 75 mL) with the last extraction occurred stirring
overnight. To give the acetal as an oil (2.32 g). The sugar was dissolved in anhydrous THF (200
mL) with BnBr (3.5 mL, 3 equiv) and TBAI (40 mg, 0.1 equiv). NaH (1.44 g 60% dispersion,
3.0 equiv) was added slowly and stirred overnight under N2. The reaction was diluted with cold
EtOAc (100 mL) and then H2O was added slowly (20 mL). The phases were separated and then
dried and concentrated. The product was purified by flash chromatography (1:9 EtOAc:Hexanes)
68
to give the dibenzylated product 7 (4.46 g, 42% over the two steps) as an oil. 1H NMR for 7
(CDCl3, 400 MHz, 295 K) δH 7.33-7.20 (m, 10H, Ar), 4.73 (s, 1H, H-1), 4.70-4.64 (m, 2H, 2x
CHHAr), 4.56-4.51 (m, 2H, 2 × CHHAr), 3.87-3.83 (m, 2H, H-2, H-3), 3.78-3.70 (m, 4H, H-4,
H-5, H-6a, H-6b) 3.34 (s, 3H, OMe), 1.53 (s, 3H, OCCH3), 1.32 (s, 3H, OCCH3). The NMR data
are in agreement with those reported in the literature.41
Methyl 4,6-di-O-benzyl-α-D-mannopyranoside (8)
Ketal 7 (3.05 g, 7.26 mmol) was dissolved in 80% AcOH (50 mL) stirred overnight at 80 oC. The
solution was co-concentrated with toluene (3 × 50 mL) and the product was purified by flash
chromatography (3:7 EtOAc:Hexanes) to give the diol as a 8 (2.71 g, 83%) clear oil. 1H NMR
for 8 (CDCl3, 400 MHz, 295 K) δH 7.33-7.20 (m, 10H, Ar), 4.73 (s, 1H, H-1), 4.70-4.64 (m, 2H,
CH2Ar), 4.56-4.51 (m, 2H, 2 × CH2Ar), 3.87 (m, 2H, H-2, H-3), 3.78 (m, 4H, H-4, H-5, H-6a, H6b) 3.34 (s, 3H, OMe). The NMR data are in agreement with those reported in the literature.40
69
Methyl 2-O-benzoyl-4,6-di-O-benzyl-α-D-mannopyranoside (3)
Diol 8 (1.84 g, 4.92 mmol) was dissolved in anhydrous MeCN (31 mL) with p-TsOH (73.6 mg,
0.09 equiv.). Trimethylorthobenzoate (1.60 mL, 1.89 equiv) was added to the reaction and was
stirred for 2.5 hours. The reaction was quenched with Et3N (1.2 mL) and concentrated to a
yellow syrup. The syrup was dissolved in 80% AcOH (100 mL) and stirred for 1 hr at RT. The
reaction was co-concentrated with toluene (3 × 50 mL) to a yellow oil. The crude was purified by
silica chromatography (4:6 EtOAc:Hex) to give the acceptor 3 (1.88 g, 80%) as an off white
solid. 1H NMR for 3 (CDCl3, 400 MHz, 295 K) δH 8.02 (d, 2H, J = 8.0 Hz, Bz), 7.57 (t, 1H, J
=8.4 Hz, Bz), 7.35-7.24 (m, 12H, Bz, Bn), 5.33 (t, 1H, J = 2.0 Hz H-2), 4.81 (s, 1H, H-1), 4.79
(d, 1H, J = 11.0 Hz, CH2Ar), 4.73 (d, 1H, J = 12.1 Hz, CH2Ar ), 4.64 (d, 1H, J = 11.1 Hz,
CH2Ar) 4.56 (d, 1H, J = 12.1 Hz, CH2Ar) 4.23 (dd, 1H, J = 3.1, 9.3 Hz, H-3), 3.99 (t, 1H, J = 9.6
Hz, H-4), 3.89 (dd, 1H, J = 4.3, 11.1, H-6a), 3.80-3.75 (m, 2H, H-5, H-6b) 3.34 (s, 3H, OMe).
The NMR data are in agreement with those reported in the literature.40
70
Methyl 3-O-(4-O-acetyl-6-anhydro-3-O-benzyl-2-O-chloroacetyl-β-D-glucopyranose)-2-Obenzoyl-4,6-di-O-benzyl-α-D-mannopyranoside (18)
Acceptor 3 (127 mg, 0.26 mmol), and donor 5 (100 mg, 0.19 mmol), was dissolved in anhydrous
DCM (2.40 mL) and stirred under N2 for 15 min at RT. A 0.01 M solution of TMSOTf was
created and added dropwise (770 μL, 0.04 equiv) to the reaction. The reaction was stirred for 6
hr and then quenched with Et3N (5 μL) and stored in the freezer overnight. The next morning the
reaction was concentrated purified by reverse-phase HPLC (CH3CN/H2O, 40:60 → 95:5) to give
the disaccharide 18 (87 mg, 54%) as a white powder. [α]D -11.5 (c 1.0, CHCl3) 1H NMR for 18
(CDCl3, 400 MHz, 295 K) δH 7.98 (d, 2H, J =7.1 Hz, Bz), 7.55 (t, 1H, J =7.5 Hz, Bz), 7.39-7.27
(m, 17H, Bn × 3, Bz), 5.39 (m, 1H, H-2), 4.99 (dd, 1H, J = 7.9, 9.5 Hz, H-2’), 4.94 (d, 1H, J = 11
Hz), 4.86 (t, 1H, J = 9.6 Hz, H-4’), 4.80 (d, 1H, J = 2 Hz, 1-H), 4.67 (d, 1H, J = 12.0 Hz
CHHAr), 4.60 (d, 1H, J = 7.9 Hz, H-1’), 4.55-4.49 (m, 4H, 4 × CHHAr), 4.42 (dd, 1H, J = 3.3,
9.3 Hz, H-3), 4.00 (t, 1H, J = 9.4 Hz, H-4), 3.90-3.81 (m, 2H, H-6A, H-5), 3.74 (dd, 1H, J = 1.4,
10.2 Hz, H-6B), 3.63 (t, 1H, J = 9.4 Hz, H-3’), 3.58-3.47 (m, 1H, H-5’), 3.36 (s, 3H, OMe), 3.24
(AB pattern, 2H, J = 12.4, 15.6 Hz, CH2Cl), 1.97 (s, 3H, OAc), 1.16 (d, 3H, J = 6.0 Hz, H-6’)
C NMR (CDCl3, 100 MHz, 295 K): δC 169.6 (C=O), 165.8 (C=O), 165.7 (C=O), 138.3, 137.7,
13
129.2 (4o Carmo), 133.6, 129.8 128.7-127.5 (Carmo), 98.3 (C-1’), 97.1 (C-1), 80.1 (C-3’), 75.6 (C3), 75.1 (CH2Ar), 74.7 (C-2’), 74.6 (C-4’), 74.2 (CH2Ar), 73.4 (CH2Ar), 72.8 (C-4), 71.3 (C-5),
70.3 (C-5’), 68.8 (C-6), 68.7 (C-2), 54.9 (OMe), 40.2 (CH2Cl), 20.8 (OAc), 17.3 (C-6’).
HRESIMS calculated for C45H49ClO13 [M+Na]+ 855.2760, found 855.2275.
71
4-O-Acetyl-6-anhydro-3-O-benzyl-1,2-O-[methyl-2-O-benzoyl-4,6-di-O-benzyl-3-Oorthochloroacetyl-α-D-mannopyranoside]-α-D-glucopyranose (19)
Acceptor 3 (74 mg, 0.15 mmol) and donor 5 (103 mg, 0.19 mmol) was dissolved in anhydrous
DCM (4 mL) with 4 Å powered molecular sieves (770 mg) while stirred under N2 for 15 min at
RT. A 1 M solution of TMSOTf was created and added dropwise (8 μL, 0.04 equiv) to the
reaction. The reaction was stirred for 1 hr and then another addition of TMSOTf was added (8
μL, 0.04 equiv). The reaction was stirred again for another hour and then another addition of
TMSOTf (8 μL, 0.04 equiv) was added. The reaction was quenched with Et3N (2 μL) and filtered
over celite. The reaction was stored in the freezer overnight. The next morning the reaction was
concentrated and purified by reverse-phase HPLC (CH3CN/H2O, 40:60 → 95:5) giving
orthoester 19 (105 mg, 85%) as a powder. [α]D 8.1 (c 0.6, MeOH) 1H NMR (CDCl3, 400 MHz,
295 K) 8.04 (d, 2H, J =7.2 Hz, Bz) 7.54 (t, 1H, J =7.6 Hz, Bz), 7.41-7.21 (m, 17H, Bn × 3, Bz),
5.76 (d, 1H, J = 5.4 Hz, H-1’), 5.46 (t, 1H, J = 2.0 Hz, H-2), 4.82-4.67 (m, 5H, 3 × CHHAr, H-1,
H-4’), 4.59-4.50 (m, 4H, 3 × CHHAr, H-2’), 4.29 (dd, 1H, J =3.2, 9.6 Hz, H-3), 4.06 (t, 1H, J
=10.0 Hz), 4.02-3.98 (m, 1H, H-5’), 3.91-3.81 (m, 4H, H-5, H-6A, H-3’, CHHCl), 3.76 (dd, 1H,
J = 1.2, 11.9 Hz, H-6B), 3.67 (d, 1H, J = 12.4 Hz, CHHCl), 3.39 (s, 3H, OMe), 2.0 (s, 3H, Ac),
1.16 (d, 3H, J = 6.4 Hz, H-6’). 13C NMR (CDCl3, 100 MHz, 295 K): δC 170.0 (C=O), 165.7
(C=O), 138.3, 137.9 137.7, 129.9 (4o Carmo), 133.3, 130.0, 128.4-128.3, 127.9-127.5 (Carom),
119.1 (OOCCCH3), 98.6 (C-1’), 98.5 (C-1), 78.7 (C-2’), 78.1 (C-3’), 75.2 (CH2Ar) 73.5 (C-5),
72
73.4 (CH2Ar), 72.6 (C-4), 72.4 (C-3), 71.9 (CH2Ar), 71.9 (C-2), 71.8 (C-4’), 68.9 (C-6), 66.9 (C5’), 55.1 (OMe), 44.3 (CH2Cl), 20.9 (Ac), 17.3 (C-6’). HRESIMS calculated for C45H49ClO13
[M+Na]+ 855.2760, found 855.2761.
Methyl 3-O-(4-O-acetyl-6-anhydro-3-O-benzyl-α-D-glucopyranose)-2-O-benzoyl-4,6-di-Obenzyl-α-D-mannopyranoside (20)
Donor 5 (50 mg, 0.096 mmol) and acceptor 3 (60 mg, 0.126 mmol) was dissolved in anhydrous
DCM (2.4 mL). The reaction was heated to 40 oC and then activated by a 0.1 M BF3•OEt2
solution (240 μL, 0.2 equiv) and was sealed. The reaction was stirred for 2 hr then cooled to RT.
The solution was quenched with Et3N (5 μL). The reaction was purified by reverse-phase HPLC
(CH3CN/H2O, 40:60 → 95:5) giving the disaccharide 18 (17.6 mg, 22%) and the title
disaccharide 20 (7.7 mg, 10%). [α]D 21.0 (c 1.0, MeOH) 1H NMR for 20 (CDCl3, 400 MHz, 295
K) δH 8.06 (d, 2H, J = 8.4 Hz, Bz), 7.58 (t, 1H, J =7.6 Hz, Bz), 7.39-7.27 (m, 17H, Bn × 3, Bz),
5.39 (dd, 1H, J = 2.0, 3.6 Hz, H-2), 5.12 (d, 1H, J = 4.0 Hz, H-1’), 4.88 (d, 1H, J = 10.4 Hz,
CHHAr), 4.84 (d, 1H, J = 2.0 Hz, H-1), 4.75 (d, 1H, J = 12.0 Hz, CHHAr), 4.72-4.48 (m, 5H, 4
× CHHAr, H-4’), 4.28 (dd, 1H, J = 3.6, 11.2 Hz, H-3), 4.19 (t, 1H, J = 9.6 Hz, H-4), 3.90 (dd,
1H, J = 3.6, 10.8 Hz, H-6A), 3.84-3.80 (m, 3H, H-5’, H-5, H-6B), 3.67 (dd, 1H, J = 4.0, 9.6 Hz,
H-2’) 3.51 (1H, t, J = 10.0 Hz, H-2’), 3.38 (s, 3H, OMe), 2.13 (d, 1H, J = 8.0 Hz, OH-2’), 1.88
(s, 3H, OC(O)CH3), 0.97 (d, 3H, J = 6.4 Hz, H-6’). 13C NMR (CDCl3, 100 MHz, 295 K): δC
73
169.9 (C=O), 165.7 (C=O), 139.3, 138.4, 138.2 (4o Carmo), 133.3, 129.9, 129.8, 128.5-127.5
(Carom), 100.1 (C-1’), 98.3 (C-1), 79.9 (C-3’), 77.4 (C-3), 74.9 (CH2Ar), 74.7 (C-4), 74.6 (C-4’),
74.3 (CH2Ar), 73.5 (CH2Ar), 72.7 (C-2’), 72.2 (C-2), 71.5 (C-5), 68.8 (C-6), 66.6 (C-5’), 55.1
(OMe), 20.9 (OAc), 17.0 (C-6’). HRESIMS calculated for C43H48O12 [M+Na]+ 779.3044, found
779.3055.
Methyl 3-O-(4-O-acetyl-6-anhydro-3-O-benzyl-β-D-glucopyranose)-2-O-benzoyl-4,6-di-Obenzyl-α-D-mannopyranoside (21)
Disaccharide 18 (377.6 mg, 0.45 mmol) was dissolved in 10 mL of a 7:3 pyridine:EtOH solution.
Thiourea (410.0 mg, 12 equiv) was added and the solution was heated to 70 oC. The reaction was
stirred for 5.5 h and then cooled. The reaction was diluted in DCM (200 mL) and was washed
with 2 M HCl (50 mL) followed by a aq NaCO3 (25 mL). The organic phase was dried over
Na2SO4 and concentrated to give the alcohol 21 (348.1 mg, quant) as a yellow foam. [α]D -23.8 (c
1.0, MeOH) 1H NMR for 21 (CDCl3, 400 MHz, 295 K) δH 8.09 (dd, 2H, J = 1.4, 8.4 Hz, Bz),
7.55 (tt, 1H, J = 1.4, 7.6 Hz, Bz), 7.39-7.23 (m, 12H, Bn × 2, Bz), 5.54 (dd, 1H, J = 2.1, 3.0 Hz,
H-2), 4.97 (d, 1H, J = 10.5 Hz, CHHAr), 4.95 (d, 1H, J = 11.6 Hz, CHHAr), 4.85 (d, 1H, J = 1.9
Hz, 1-H), 4.78-4.67 (m, 3H, H-4’, 2 × CHHAr), 4.55 (d, 1H, J = 11.6 Hz, CHHAr), 4.48 (d, 1H,
J = 7.4 Hz, H-1’), 4.42 (d, 1H, J = 10.4 Hz, CHHAr), 4.34 (dd, 1H, J = 2.9, 9.9 Hz, H-3), 4.05 (t,
1H, J = 9.7 Hz, H-4), 3.90 (dd, 1H, J = 3.6, 10.6 Hz, H-6A), 3.86-3.83 (m, 1H, H-5), 3.79 (dd,
74
1H, J = 1.3, 10.3 Hz, H-6B), 3.64 (bs, 1H, OH-2), 3.53-3.50 (m, 2H, H-2’, H-3’), 3.43- 3.39 (m,
4H, H-5’, OMe), 1.93 (OAc), 1.04 (d, 3H, J = 6.1 Hz, H-6’). 13C NMR (CDCl3, 100 MHz, 295
K): δC 170.0 (C=O), 167.0 (C=O), 138.7, 138.4, 138.3, 129.3 (4o Ar), 133.7, 130.2 128.6-127.4
(Ar), 103.7 (C-1’), 98.7 (C-1), 81.4 (C-3), 79.8 (C-3’), 75.1 (CH2Ar), 75.0 (C-4), 74.4 (C-4’),
74.3 (CH2Ar), 73.5 (C-2’), 73.4 (CH2Ar), 71.7 (C-5), 71.1 (C-2), 70.2 (C-5’), 68.9 (C-6), 55.0
(OMe), 20.9 (OAc), 17.4 (C-6’). HRESIMS calculated for C43H48O12 [M+Na]+ 779.3044, found
779.3045.
Methyl 3-O-(4-6-dianhydro-3-O-benzyl-2-oxo-α-D-erythro-hex-3-enopyranoside)-2-Obenzoyl-4,6-di-O-benzyl-α-D-mannopyranoside (24)
In a 2 mL vial alcohol 21 (100 mg, 0.132 mmol) was sealed and transferred into a glove box.
Anhydrous DMSO (750 μL) was added followed by Ac2O (377 μL, 26.8 equiv) and the reaction
was allowed to stir for 18 hr at room temperature. The reaction was then diluted with EtOAc (5
mL) and washed with brine (1 mL) and water (3 × 1 mL). The organic phase was dried over
sodium sulfate and concentrated to an yellow oil. The crude was purified by silica
chromatography (3:7 EtOAc:Hex) to give eliminated 24 (39.7 mg, 43%) as well as the oxidized
23 (8.2 mg, 8%) as oils. [α]D 3.500 (c 1.0, MeOH)1H NMR for 24 (CDCl3, 400 MHz, 295 K) δH
8.10 (d, 2H, J = 8.4 Hz, Bz), 7.60 (t, 1H, J =7.6 Hz, Bz), 7.39-7.27 (m, 12H, Bn × 2, Bz), 5.76
(d, 1H, J = 2.5 Hz, H-4’), 5.56 (dd, 1H, J = 2.0, 3.3 Hz, H-2), 5.18 (s, 1H, H-1’), 4.92 (d, 1H, J =
75
10.5 Hz, CHHAr), 4.88 (d, 1H, J = 12.2 Hz, CHHAr), 4.87 (d, 1H, J = 1.9 Hz, H-1), 4.79 (d, 1H,
J = 12.1 Hz, CHHAr), 4.72 (d, 1H, J = 12.0 Hz, CHHAr), 4.66-4.60 (m, 1H, H-5’), 4.55-4.47 (m,
3H, 2 × CHHAr, H-3), 4.12 (t, 1H, J = 9.6 Hz, H-4), 3.89-3.82 (m, 2H, H-5, H-6A), 3.78 (bd,
1H, J = 8.7 Hz), 3.39 (s, 3H, OMe), 1.36 (d, 3H, J = 6.8 Hz, H-6’). 13C NMR (CDCl3, 100 MHz,
295 K): δC 185.0 (C-2’), 165.8 (C=O), 147.0 (C-3’), 138.2, 135.6, 133.3, 129.9, 129.7, 128.5127.4 (Carom), 120.5 (C-4’), 98.5 (C-1), 96.6 (C-1’), 75.9 (C-3), 74.9 (CH2Ar), 73.4 (C-4), 73.3
(CH2Ar), 71.1 (C-5), 69.8 (CH2Ar), 68.7 (C-6), 68.7 (C-5’), 54.9 (OMe), 22.3 (C-6’). HRESIMS
calculated for C41H42O10 [M+Na]+ 717.2670, found 717.2639.
Methyl 3-O-(4-O-acetyl-6-anhydro-3-O-benzyl-β-D-mannopyranose)-2-O-benzoyl-4,6-diO-benzyl-α-D-mannopyranoside (22)
In a 2 mL vial alcohol 21 (48 mg, 0.064 mmol) was sealed and transferred into a glove box.
Anhydrous DMSO (360 μL) was added followed by Ac2O (90 μL, 13.4 equiv) and the reaction
was allowed to stir for 6 hr at room temperature. The reaction was then diluted with EtOAc (5
mL) and washed with brine (1 mL) and water (3 × 1 mL). The organic phase was dried over
sodium sulfate and concentrated to an yellow oil. The oil was dissolved in freshly distilled THF
(1 mL) and cooled and stirred at 78 oC for 15 min. L-Selectride® (1 M in THF, 250 μL, 4 equiv)
was added dropwise and the reaction was allowed to stir for 20 min. The reaction was quenched
with AcOH (250 μL) and then warmed to RT. The solution was concentrated and purified by
76
reverse-phase HPLC (CH3CN/H2O, 40:60 → 95:5) to give the inverted 22 (22 mg, 44%) as an
oil. [α]D -37.3 (c 1.0, MeOH) 1H NMR for 22 (CDCl3, 400 MHz, 295 K) δH 8.02 (d, 2H, J = 7.7
Hz, Bz), 7.58 (t, 1H, J =7.7 Hz, Bz), 7.39-7.21 (m, 17H, Bn × 3, Bz) 5.49 (dd, 1H, J = 2.0, 3.3
Hz, H-2), 5.05 (t, 1H, J = 9.6 Hz, H-4’), 4.95 (d, 1H, J = 10.2 Hz, CHHAr), 4.83 (d, 1H, J = 2.2
Hz, H-1), 4.74 (d, 1H, J = 12.0 Hz, CHHAr), 4.67 (d, 1H, J = 12.3 Hz, CHHAr), 4.67 (bs, 1H,
H-1’), 4.55 (d, 1H, J = 12.0 Hz, CHHAr), 4.50-4.44 (m, 3H, 2 × CHHAr, H-3), 4.02 (t, 1H, J =
9.5 Hz, H-4), 3.96 (bd, 1H, J = 1.2 Hz, H-2’), 3.89 (dd, 1H, J = 3.9, 10.4 Hz, H-6A), 3.84 (m,
1H, H-5), 3.80 (dd, 1H, J = 1.7, 10.4 Hz, H-6A), 3.46 (dd, 1H, J = 3.1, 9.6 Hz, H-3’), 3.40-3.36
(m, 1H, H-5’), 3.38 (s, 3H, OMe), 2.00 (s, 3H, OAc), 1.19 (d, 3H, J = 6.2 Hz, H-6’) 13C NMR
(CDCl3, 100 MHz, 295 K): 170.0 (C=O), 166.0 (C=O), 138.3, 138.2 137.7, 129.5 (4o Carmo),
133.4, 129.9, 128.5-127.5 (Carmo), 98.7 (C-1’), 96.8 (C-1), 78.5 (C-3’), 75.8 (C-3), 75.0 (CH2Ar),
73.4 (CH2Ar), 73.0 (C-4), 72.2 (C-4’), 71.2 (C-5), 71.0 (CH2Ar), 70.5 (C-5’), 68.9 (C-6), 68.7
(C-2’), 68.0 (C-2), 54.9 (OMe), 21.0 (OAc), 17.5 (C-6’). HRESIMS calculated for C43H48O12
[M+Na]+ 779.3038, found 779.3002.
Methyl 3-O-(4-O-acetyl-6-anhydro-2-O-benzoyl-3-O-benzyl-β-D-mannopyranose)-2-Obenzoyl-4,6-di-O-benzyl-α-D-mannopyranoside (25)
Sugar 22 (31 mg, 0.041 mmol) was dissolved in anhydrous 1,2 dichloroethane (1 mL) and
anhydrous pyridine (30 μL, 8 equiv). BzCl (15 μL, 3 equiv) was added and the reaction was
sealed and heated to 40 oC. The reaction was stirred for 15 hrs and then allowed to cool. The
77
reaction was diluted with DCM (5 mL) and then washed with 1 M HCl. The organic phase was
dried and concentrated. The crude was purified by silica chromatography (2:8 EtOAc:Hex) to
give the benzoylated 25 (32 mg, 91%) as an oil. [α]D -49.8 (c 1.0, MeOH) 1H NMR for 25
(CDCl3, 400 MHz, 295 K) 8.07 (dd, 2H, J = 1.4, 7.1 Hz, Bz), 7.79 (dd, 2H, J = 1.4, 8.4 Hz, Bz),
7.58 (t, 1H, J = 8.8 Hz, Bz), 7.41 (t, 2H, J = 8.1 Hz, Bz), 7.35-7.06 (m, 18H, Bn × 3, Bz), 5.70
(bd, 1H, J = 2.3 Hz, H-2’), 5.48 (dd, 1H, J = 1.9, 3.3 Hz, H-2), 5.04 (t, 1H, J = 9.7 Hz, H-4’),
4.86 (bs, 1H, H-1), 4.86 (d, 1H, J = 2.2 Hz, H-1’), 4.75 (d, 1H, J = 10.4 Hz, CHHAr), 4.70 (d,
1H, J = 12.2 Hz, CHHAr), 4.64 (d, 1H, J = 12.0 Hz, CHHAr), 4.49-4.43 (m, 3H, 2 × CHHAr, H3), 4.04 (d, 1H, J = 10.4 Hz, CHHAr), 3.79-3.67 (m, 4H, H-4, H-5, H-6AB), 3.63 (dd, 1H, J =
3.3, 9.8 Hz, H-3’), 3.53 (m, 1H, H-5’), 3.37 (s, 1H, OMe), 2.00 (s, 3H, OAc), 1.31 (d, 3H, J =6.2
Hz). 13C NMR (CDCl3, 100 MHz, 295 K): 170.0 (C=O), 166.3 (C=O), 165.3 (C=O), 138.4,
138.3, 137.7, 129.7, 129.6 (4o Carmo), 130.0, 129.7, 128.4-127.8 (Carmo), 98.6 (C-1), 95.5 (C-1’),
76.5 (C-3’), 76.0 (C-3), 74.8 (CH2Ar), 73.4 (CH2Ar), 72.8 (C-4’), 72.5 (C-4), 71.0 (C-5’), 70.6
(C-5), 70.6 (CH2Ar), 68.9 (C-6), 68.4 (C-2’), 68.0 (C-2), 54.9 (OMe), 21.0 (Ac), 17.8 (C-6’).
HRESIMS calculated for C50H52O13 [M+Na]+ 883.3300, found 889.3287.
78
Methyl 3-O-(6-anhydro-2-O-benzoyl-3-O-benzyl-β-D-mannopyranose)-2-O-benzoyl-4,6-diO-benzyl-α-D-mannopyranoside (26)
Protected disaccharide 25 (29 mg, 0.033 mmol) was dissolved in anhydrous MeOH (1 mL).
AcCl (30 μL, 0.42 mmol) was added dropwise to the reaction. The reaction was heated to 50 oC
and was allowed to stir for 4 hr. The solution was then cooled to RT and then quenched with
Et3N (100 μL). The reaction was concentrated and then purified by column chromatography (2:8
EtOAc:Hex) giving the disaccharide acceptor 26 (21.8 mg, 81%) as an oil. [α]D -32.9 (c 1.0,
MeOH) 1H NMR for 26 (CDCl3, 400 MHz, 295 K) δH 8.03 (dd, 2H, J = 1.4, 8.5 Hz, Bz), 7.74
(dd, 2H, J = 1.2, 8.4 Hz, Bz), 7.53 (tt, 1H, J = 2.5, 7.7 Hz, Bz), 7.38 (bt, 2H, J = 8.3 Hz, Bz),
7.32-7.08 (m, 16H, Bn × 3, Bz), 7.04 (bt, 2H, J = 7.7 Hz, Bz), 5.67 (bd, 1H, J = 2.1 Hz, H-2’),
5.47 (dd, 1H, J = 2.0, 3.3 Hz, H-2), 4.87 (bs, 1H, J = 0.8 Hz, 1-H’), 4.83 (bd, 1H, 1.9 Hz, 1-H),
4.80 (d, 1H, J = 11.1 Hz, CHHAr), 4.76 (d, 1H, J = 10.4 Hz, CHHAr), 4.62 (d, 1H, J = 12.1 Hz,
CHHAr), 4.46-4.43 (m, 2H, CHHAr, H-3), 4.39 (d, 1H, J = 11.1 Hz, CHHAr), 4.04 (d, 1H, J =
10.4 Hz, CHHAr), 3.77-3.75 (m, 2H, H-4, H-5), 3.73 (dd, 1H, J = 2.8, 10.8 Hz, H-6A), 3.66 (bd,
1H, J = 9.4 Hz, H-6B), 3.56 (bt, 1H, J = 9.8 Hz, H-4’), 3.48 (dd, 1H, J = 3.0, 9.6 Hz), 3.45-3.41
(m, 1H, H-5) 3.35 (s, 3H, OMe), 2.24 (bs, 1H, OH-4), 1.42 (d, 3H, J = 6.0 Hz, H-6’). 13C NMR
(CDCl3, 100 MHz, 295 K): δC 166.2 (C=O), 165.2 (C=O), 138.4, 138.3 137.3, 129.7 (4o Carmo),
133.1, 132.6, 129.9, 129.6, 128.6-127.9 (Carmo), 98.6 (C-1’), 95.5 (C-1), 79.8 (C-3’), 75.8 (C-3),
74.7 (CH2Ar), 73.3 (CH2Ar), 72.8 (C-4), 72.0 (C-5’), 71.8 (C-4’), 71.0 (CH2Ar, C-5), 68.9 (C-2),
79
67.8 (C-2’), 54.9 (OMe), 18.0 (C-6’). HRESIMS calculated for C48H50O12 [M+Na]+ 841.3194,
found 841.3160.
Methyl 3-O-[6-anhydro-4-O-(2,3,4,6-tetra-O-benzoyl-α-D-mannopyranoside)-2-O-benzoyl3-O-benzyl-β-D-mannopyranose]-2-O-benzoyl-4,6-di-O-benzyl-α-D-mannopyranoside (27)
Disaccharide acceptor 26 (9.1 mg, 0.011 mmol) and mannosyl donor (18.9 mg, 2.5 equiv) were
dissolved in anhydrous DCM (500 μL) with 4 Å molecular sieves (50 mg). The reaction was
cooled and stirred at -40 oC for 30 min. NIS (8.3 mg, 3 equiv) was added followed by the
dropwise addition of a 0.01 M TfOH solution (80 μL, 0.07 equiv) and was allowed to warm to 0
C over 2 hr. The reaction was quenched with Et3N (2 μL) and diluted with DCM (5 mL). The
o
reaction was filtered through Celite and the filtrate was washed aq. Na2S2O3 (5 mL). The organic
phase was dried over Na2SO4 and concentrated. The crude was purified by silica chromatograph
(1:9 EtOAc:Hex) to give trisaccharide 27 (3.4 mg, 19%) as an oil. 1H NMR for 27 (CDCl3, 400
MHz, 295 K) δH 8.07 (dd, 2H, J = 1.2, 8.5 Hz, Bz), 7.91 (bt, 1H, J = 8.5 Hz, Bz), 7.81 (dd, 2H, J
= 1.4, 8.5 Hz, Bz), 7.73 (dd, 2H, J = 1.2, 8.4 Hz, Bz), 7.48-7.13 (m, 31H, Bn × 3, Bz), 7.05-6.98
(m, 7H, Bn × 3, Bz), 6.00 (t, 1H, J = 9.9 Hz, H-4’’), 5.82-5.79 (m, 2H, H-2’’, H-3’’), 5.74 (bs,
1H, H-2’), 5.54-5.52 (m, 2H, H-2, H-1’’), 4.90 (bs. 1H, H-1’), 4.88 (d, 1H, J = 1.7 Hz, H-1),
4.76 (d, 1H, J = 10.6 Hz, CHHAr), 4.72 (d, 1H, J = 10.6 Hz, CHHAr), 4.67-4.62 (m, 2H,
CHHAr, H-6A’’), 4.52-4.43 (m, 5H, 2 × CHHAr, H-3, H-5’’, H-6B’’), 4.05 (d, 1H, J = 10.7 Hz,
80
CHHAr), 3.80-3.63 (m, 7H, H-4, H-5, H-6AB, H-3’, H-4’, H-5’), 3.40 (s, 3H, OMe), 1.54 (d,
3H, J = 6.1 Hz, H-6’) 13C NMR (CDCl3, 100 MHz, 295 K): δC 166.2, 165.5, 165.4, 165.1, 164.8
(C=O), 138.3, 136.5, 129.3, 129.1, 128.9 (4o Carmo), 133.4-132.6, 130.0-127.4 (Carmo), 99.6 (C1’’), 98.7 (C-1), 95.4 (C-1’), 79.5 (C-3’), 79.2 (C-3’’), 76.1 (C-3), 74.7 (CH2Ar), 73.4 (CH2Ar),
72.9 (C-4), 71.0 (C-4’), 70.8 (CH2Ar), 70.1 (C-2’’, C-5), 69.7 (C-5’, C-5’’), 68.9 (C-6), 68.3 (C2, C-2’), 67.8 (C-4’’), 63.7 (C-6’’), 54.9 (OMe), 19.0 (C-6). HRESIMS calculated for C82H76O21
[M+Na]+ 1419.4771, found 1419.4797.
S-Phenyl 2,3-O-carbonyl-1-thio-α-L-rhamnopyranoside (30)
Triol 29 (1.20 g, 3.24 mmol) was dissolved in pyridine (24 mL) and cooled to 0 °C. Slowly a
15% phosgene in toluene solution (3.56 ml, 1.6 eq) was added dropwise and the reaction was
stirred for 3h. The reaction was quenched with the slow addition of ice cold saturated NaHCO3
(50 mL) and then diluted with ethyl acetate (50 mL) and separated. The water phase was reextracted with EtOAc and the combined organic phase was washed with 1N HCl (400 mL),
saturated NaHCO3 (50 mL) and was dried over sodium sulfate and concentrated. The crude
product was triturated with Et2O to give known carbonate 30 (557.3 mg, 61%) white solid. 1H
NMR for 30 (CDCl3, 400 MHz, 295 K) δH 7.20-7.55 (m, 5H, Ar), 5.78 (s, 1H, H-1), 4.88 (d, 1H,
81
J = 6.7 Hz, H-2), 4.71 (t, 1H, J =7.0 Hz, H-3), 4.12-4.26 (m, 1H, H-4), 3.56-3.68 (m, 1H, H-5),
2.48 (d, 1H, J = 4.7 Hz, OH-4), 1.23 (d, 3H, J = 6.1 Hz, H-6). The NMR data are in agreement
with those reported in the literature.54
S-Phenyl 2,3-O-carbonyl-4-O-chloroacetyl-1-thio-α-L-rhamnopyranoside (31)
The alcohol 30 (1.00 g, 3.67 mmol) was dissolved in anhydrous DCM (50 mL) and anhydrous
pyridine (1 mL, 3.4 equiv). The reaction nwas cooled to 0 oC and ClAcCl (438 μL, 1.5 equiv)
was added dropwise to the solution. The solution was stirred for 1 hr and then was washed with 1
M HCl (14 mL) followed by sat’d NaHCO3 (10 mL) wash. The organic phase was dried and
concentrated to a solid. The crude was triturated with ether to give the chloroacetyl 31 (1.12 g,
88%) as an off white powder. [α]D -110.6 (c 1.0, CH2Cl2) 1H NMR for 31 (CDCl3, 400 MHz, 295
K) 7.42 (m, 2H, SAr), 7.35 (m, 3H, SAr), 5.79 (s, 1H, H-1), 5.03 (dd, 1H, J = 7.1 Hz, 9.4 Hz, H4), 4.92 (d, 1H, J = 6.9 Hz, H-3), 4.83 (t, 1H, J = 7.0 Hz, H-2), 4.37-4.34 (m, 1H, H-5), 4.13 (s,
2H, CH2Cl), 1.20 (d, 3H, J = 6.3 Hz, H-6). 13C NMR (CDCl3, 100 MHz, 295 K): δC 166.0
(ClAc), 152.5 (OCOCO), 132.5 (SAr), 131.2 (SAr), 129.5 (SAr), 128.7 (SAr), 82.3 (C-1 J =
166.6 Hz), 77.1 (C-3), 75.5 (C-4), 74.6 (C-2), 64.9 (C-5), 40.4 (ClAc), 17.1 (C-6). HREI-MS
calculated for C15H15O6ClS [M]+ 358.0278, found 358.0278.
82
2,3-O-Carbonyl-4-O-chloroacetyl-α-L-rhamnopyranoside bromide (32)
Thiorhamnoside 31 (0.52 g, 1.45 mmol, 1 eq) was dissolved in anhydrous DCM (10 mL) was
treated with elemental bromine (156 μL, 2.1 eq) at 0 °C for 2h. The reaction was quenched with
1-hexene (300 μL) and the organic phase was washed with water and then dried and
concentrated. The crude product was triturated with hexane to afford the bromide 32 (470 mg,
99%) as an orange solid. 1H NMR for 32 (CDCl3, 400 MHz, 295 K) δH 6.59 (s, 1H, H-1), 5.07
(d, 1H, J = 6.1 Hz, H-2), 5.02 (q, 1H, J = 3.5 Hz, 7.5 Hz, H-4), 4.93 (t, 1H, J = 7.2 Hz, H-3),
4.17-4.08 (m, 1H, H-5), 4.11 (s, 2H, CH2Cl), 1.29 (d, J = 6.3 Hz, H-6) 13C NMR (CDCl3, 100
MHz, 295 K): δC 165.9 (ClAc), 151.9 (OOC=O), 80.8 (C-1), 79.0 (C-2), 74.6 (C-3), 74.1 (C-4),
68.5 (C-5), 40.3 (ClAc), 16.8 (C-6).
83
Methyl 3-O-(2, 3-O-carbonyl-4-O-chloroacetyl-α-L-rhamnopyranoside)- 2-O-benzoyl-4,6di-O-benzyl-α-D-mannopyranoside (34)
Acceptor 4 (50 mg, 0.10 mmol), bromide 32 (51 mg, 0.15 mmol), Ag2SiO3 (72 mg, 2 equiv) and
AgOTf (4 mg, 0.1 equiv) were suspended in DCM (5 ml) with 4 Å molecular sieves (50 mg) and
placed under N2. The reaction was stirred vigorously at RT for 3 hr. After it was diluted in DCM
(10 mL) and was filtered over Celite. The filtrate was washed with sat’d NaHCO3 (10 mL) and
the organic phase was dried and concentrated. The crude was purified by silica chromatography
(1:9 EtOAc:Tol) but was unable to get pure. The yield of disaccharide 34 is estimated by NMR
(18 mg, 25%) as an oil. 1H NMR for 34 (CDCl3, 600 MHz, 295 K) δH 8.05 (d, 2H, J = 7.6 Hz,
Bz), 7.58 (t, 1H, J = 7.4 Hz, Bz), 7.39-7.24 (m, 10H, Bn, Bz), 7.16 (d, 2H, J = 5.8 Hz, Bz), 5.52
(bs, 1H, H-2), 5.40 (s, 1H, H-1’), 4.85 (s, 1H, H-1), 4.80-4.78 (m, 2H, H-2’, CHHAr), 4.63 (m,
3H, 3 × CHHAr), 4.47 (t, 1H, J = 7.1 Hz, H-4’), 4.35 (d, 1H, J = 7.1 Hz, H-3’), 4.26 (dd, 1H, J =
2.8, 9.6 Hz, H-3), 4.03 (t, 1H, J = 9.4 Hz, H-4), 3.99-3.77 (m, 6H, H-5’, H-5, H-6A,B, CH2Cl),
3.41 (s, 3H, OMe), 0.93 (d, 3H, J = 6.5 Hz, H-6’) 13C NMR (CDCl3, 150 MHz, 295 K): δC 165.8
(C=O), 165.5 (C=O), 158.5 (OOC=O), 138.2, 137.8, 129.3 (4O Ar), 133.5, 130.0, 128.6-127.3
84
(Ar), 98.9 (C-1 J = 173.6 Hz), 91.4 (C-1’ J = 167.2 Hz), 76.1 (C-3’), 75.8 (C-4’), 75.0 (CH2Ar),
74.7 (C-2’), 73.5 (CH2Ar), 73.4 (C-3), 71.8 (C-5’), 68.7 (C-6), 67.8 (C-2), 62.7 (C-5), 40.2
(CH2Cl), 16.7 (C-6’). HRESIMS calculated for C37H39O13Cl [M+Na]+ 749.1982, found
749.1978.
Allyl 2,3-di-O-isopropylidene-α-L-rhamnopyranoside (35)
L-Rhamnose monohydrate (10.01 g, 60.9 mmol) was suspended in allyl alcohol (80 mL) and
then triflic acid (800 μL) was added and the reaction was refluxed for 5 hours. The reaction was
then cooled to RT and quenched with Et3N (2 mL). The reaction was concentrated to a yellow oil
and then dissolved in anhydrous acetone (60 mL) and DMP (15 mL). p-TsOH (101 mg) was
added and then placed under nitrogen and stirred overnight. The solution was quenched with
Et3N (500 μL) and concentrated. The product was purified by silica chromatography (3:7
EtOAc:Hexanes) to give the known acetal 36 (10.69 g, 72%) as a slight yellow oil. 1H NMR
(CDCl3, 400 MHz, 295 K): 5.89–5.86 (m, 1H, CH=), 5.29–5.17 (m, 2H, CH2=), 4.96 (s, 1H, H1), 4.30–4.17 (m, 2H, H-2, OCH2=), 4.06 (t, 1H, J = 7.1 Hz, H-3), 4.02–3.97 (m, 1H, OCH2=),
3.74–3.65 (m, 1H, H-5), 3.38–3.43 (m, 1H, J = 5.4, 9.3 Hz, H-4), 2.64 (d, 1H, J = 5.4, OH-4),
85
1.53 (s, 3H, CH3), 1.36 (s, 3H, CH3), 1.29 (d, 3H, J = 6.6 Hz, H-6). The NMR data are in
agreement with those reported in the literature.57
Allyl 2,3-O-isopropylidene-4-O-levulinoyl-α-L-rhamnopyranoside (36)
Sugar 35 (3.95 g, 16.1 mmol) was dissolved in anhydrous pyridine (15 mL). Levulinic anhydride
(18.77 g, 5.4 equiv) and DMAP (401 mg, 0.2 equiv) was added and the reaction was allowed to
stir overnight under N2. The next day the solution was diluted with DCM (100 mL) and then
washed with 1 M HCl (200 mL) followed by sat’d NaHCO3 (50 mL). The organic phase was
dried and concentrated and was purified by silica chromatography (1:9 to 3:7 EtOAc:Hexanes)
the levulinoate 36 (4.89 g 89%) as a yellow oil. [α]D -19.0 (c 1.0, MeOH) 1H NMR for 36
(CDCl3, 400 MHz, 295 K) δH 5.93-5.83 (m, 1H, OCH2CH=CH2), 5.31-5.19 (m, 2H,
OCH2CH=CH2), 5.03 (s, 1H, H-1), 4.83 (dd, 1H, 7.1 Hz, 10.1 Hz, H-4), 4.20-4.12 (m, 3H, H-2,
H-3, OCH2CH=CH2), 4.01-3.96 (m, 1H, OCH2CH=CH2), 3.77-3.70 (m, 1H, H-5), 2.89-2.66 (m,
2H, CH3COCH2CH2CO), 2.64-2.49 (m, 2H, CH3COCH2CH2CO), 2.16 (s, 3H,
CH3COCH2CH2CO), 1.54 (s, 3H, OCCH3), 1.31 (s, 3H, OCCH3), 1.17 (d, 3H, J = 6.3 Hz, H-6).
C NMR (CDCl3, 100 MHz, 295 K): δC 206.4 (CH3COCH2CH2CO), 172.0
13
(CH3COCH2CH2CO), 133.5 (OCH2CH=CH2), 117.9 (OCH2CH=CH2), 109.7 (OOC(CH3)2) 96.0
(C-1), 75.9 (C-3), 75.7 (C-2), 74.8 (C-4), 68.1 (OCH2CH=CH2), 64.0 (C-5), 37.9, 27.9
(COCH2CH2CO), 29.8 (CH3COCH2CH2CO), 27.7, 26.4 (OOC(CH3)2), 16.9 (C-6). HRESIMS
calculated for C43H48O12 [M+Na]+ 365.1570, found 365.1553.
86
Allyl 4-O-levulinoyl-α-L-rhamnopyranoside (37)
Acetal 36 (3.57g, 10.4 mmol) was dissolved in 80% AcOH (100 mL) and heated to 80 oC. The
reaction was stirred for 4 hrs and then co-concentrated with toluene (3 × 75 mL). Diol 37 (3.59 g
89% ) appeared as a white powered which was purified by triturated with hexanes. [α]D -46.4 (c
1.0, MeOH) 1H NMR for 37 (CDCl3, 400 MHz, 295 K) δH 5.94-5.98 (m, 1H, OCH2CH=CH2),
5.33-5.20 (m, 2H, OCH2CH=CH2), 4.90-4.84 (m, 2H, H-1, H-4), 4.20-4.14 (m, 3H, H-2, H-3,
OCH2CH=CH2), 4.04-3.91 (m, 3H, H-2, H-3, OCH2CH=CH2), 3.84-3.78 (m, 1H, H-5), 2.842.78 (m, 2H, CH3COCH2CH2CO), 2.58-2.32 (m, 2H, CH3COCH2CH2CO), 2.18 (s, 3H,
CH3COCH2CH2CO), 1.22 (d, 3H, J = 6.3 Hz, H-6). 13C NMR (CDCl3, 100 MHz, 295 K): δC
207.5 (H3CC=O), 173.4 (OC=O), 133.6 (OCH2CH=CH2), 117.5 (OCH2CH=CH2), 98.3 (C-1),
75.5 (C-4), 70.8 (C-2), 70.1 (C-3), 68.1 (OCH2CH=CH2), 65.5 (C-5), 38.3, 28.2
(COCH2CH2CO), 29.8 (CH3COCH2CH2CO), 17.3 (C-6). HRESIMS calculated for C14H22O7 [MH2O+H]+, 285.1332 found 285.1330.
87
Allyl 3-O-benzyl-4-O-levulinoyl-α-L-rhamnopyranoside (39)
Diol 37 (293 mg, 0.97 mmol) was dissolved in anhydrous toluene (20 mL) with dibutyl tin oxide
(250 mg, 1 mmol). The reaction was reflux in a Deen-Stark apparatus for 3 hr. The reaction was
cooled and concentrated to a foam. The crude was dissolved in DMF (20 mL) with CsF (295 mg,
2 equiv) and BnBr (117 μL, 1.01 equiv). The reaction was stirred overnight at RT. The next
morning the reaction was concentrated and then diluted in EtOAc (50 mL). The reaction was
washed with water (3 × 5 mL). The organic phase was dried over Na2SO4 and concentrated The
crude was purified by silica chromatography (1:8 →1:9 EtOAc:Hex) to give the alcohol product
38 (353.7 mg, 93%) as an yellow oil. [α]D -33.8 (c 1.0, MeOH) 1H NMR for 38 (CDCl3, 400
MHz, 295 K) 7.37-7.25 (m, 5H, Bn), 5.93-5.82 (m, 1H, CH2CH=CH2), 5.29-5.27 (m, 1H,
CH2CH=CH2), 5.21 5.06 (CH2CH=CH2), 5.27 (t, 1H, J = 9.7 Hz, H-4), 4.84 (d, 1H, J = 1.6 Hz,
H-1), 4.63 (d, 1H, J = 12.0 Hz, CH2Ph), 4.54 (d, 1H, J = 12.0 Hz, CH2Ph), 4.18-4.12 (m, 1H,
CH2CH=CH2), 4.01 (bs, 1H, H-2), 3.98-3.94 (m, 1H, CH2CH=CH2), 3.72-3.70 (m, 2H, H-3, H5), 2.80-2.38 (m, 5H, CH3COCH2CH2CO, OH-2), 2.13 (s, 3H, CH3COCH2CH2CO), 1.17 (d, 3H,
J = 6.3 Hz, H-6). 13C NMR (CDCl3, 100 MHz, 295 K): δC 206.4 (CH3COCH2CH2CO), 172.0
(CH3COCH2CH2CO), 137.7 (4o Ar), 133.6 (OCH2CH=CH2), 128.5, 127.9, 127.7 (C Ar), 117.5
(OCH2CH=CH2), 98.1 (C-1), 76.7 (C-3), 72.7 (C-4), 71.8 (CH2Ph), 68.4 (C-2), 68.0
88
(OCH2CH=CH2), 66.1 (C-5), 37.8, 28.0 (CH3COCH2CH2CO), 29.9 (CH3COCH2CH2CO), 17.3
(C-6). HRESIMS calculated for C21H25O6 [M-H2O-H]-, 373.1656 found 373.1652.
Allyl 3-O-benzyl-4-O-levulinoyl-2-O-trifyl-α-L-rhamnopyranoside (39)
Alcohol 38 (361 mg) was dissolved in anhydrous DCM (10 mL) and anhydrous pyridine (600
μL, 7 equiv). The reaction was sealed under N2 and cooled to -20 oC. Tf2O (310 μL, 2 equiv) was
added drop wise to the reaction. The reaction was allowed to warm to 0 oC over 45 min. The
reaction was diluted in DCM (10mL) and was washed with 1N HCl (10 mL) followed by sat’d
NaHCO3 (10 mL). The organic phase was dried over Na2SO4 and concentrated to give triflate 39
(458 mg, 95%) as a clear oil. 1H NMR for 39 (CDCl3, 400 MHz, 295 K) δH 7.39-7.26 (m, 5H,
Bn), 5.89-5.78 (m, 1H, CH2CH=CH2), 5.28-5.22 (m, 2H, CH2CH=CH2), 5.05 (bs, 1H, H-2), 4.98
(t, 1H, J = 9.8 Hz, H-4), 4.95 (d, 1H, J = 1.6 Hz, H-1), 4.74 (d, 1H, J = 12.1 Hz, CHHAr), 4.45
(d, 1H, J = 12.1 Hz, CHHAr), 4.16 (dd, 1H, J = 5.2, 12.9 Hz, CHHCH=CH2), 3.99 (dd, 1H, J =
5.2, 12.9 Hz, CHHCH=CH2), 3.89 (dd, 1H, J = 3.0, 9.7 Hz, H-3), 3.78-3.74 (m, 1H, H-5), 2.792.29 (m, 4H, CH3COCH2CH2CO), 2.13 (s, 3H, CH3COCH2CH2CO), 1.17 (d, 3H, J = 6.3 Hz, H6). 13C NMR (CDCl3, 100 MHz, 295 K): δC 206.4 (CH3COCH2CH2CO), 171.6
(CH3COCH2CH2CO), 137.1 (4o Ar), 132.6 (OCH2CH=CH2), 128.6, 127.9, 127.1 (Ar), 120.6
89
(CF3), 118.5 (OCH2CH=CH2), 95.7 (C-1), 82.0 (C-2), 73.6 (C-4), 72.3 (CH2Ar), 71.9 (C-3), 68.5
(OCH2CH=CH2), 37.8, 27.8 (COCH2CH2CO), 29.8 (CH3COCH2CH2CO), 17.2 (C-6).
Allyl 2-6-dianhydro-3-O-benzyl-4-O-levulinoyl-α-L-erythro-hex-3-enopyranoside (41)
The triflate 39 (51 mg, 0.097 mmol) was dissolved in anhydrous DMF (2 mL). Potassium acetate
(95 mg, 10 equiv) was added and the reaction flask was sealed and stirred at 40 oC for 6 hr. The
reaction was then diluted with EtOAc (10 mL) and washed with water (4 × 2 mL). The organic
phase was dried over Na2SO4 and concentrated. The crude was purified by silica chromatography
(2:8 EtOAc:Hex) to give the eliminated product 41 (36.2 mg, 88%) as an yellow oil. [α]D 51.0 (c
1.0, MeOH) 1H NMR for 41 (CDCl3, 400 MHz, 295 K) δH 7.33-7.25 (m, 5H, Ar), 5.91-5.87 (m,
1H, OCH2CH=CH2), 5.30-5.16 (m, 3H, H-4, OCH2CH=CH2), 5.16 (d, 1H, J = 2.7 Hz, H-2), 4.87
(d, 1H, J = 2.7 Hz, H-1), 4.76 (d, J = 11.7 Hz, CH2Ar), 4.71 (d, J = 11.7 Hz, CH2Ar), 4.21 (bdd,
1H, J = 5.3, 12.7 Hz, OCHHCH=CH2), 4.16-4.12 (m, 1H, H-5), 4.03 (bdd, 1H, = 6.3, 12.7 Hz,
OCHHCH=CH2), 2.71-2.45 (m, 4H, CH3COCH2CH2CO), 2.08 (s, 3H, CH3COCH2CH2CO),
1.23 (d, 3H, J = 6.3 Hz, H-6). 13C NMR (CDCl3, 100 MHz, 295 K): δC 206.3
(CH3COCH2CH2CO), 172.4 (CH3COCH2CH2CO), 154.3 (C-3), 136.2 (4o Ar), 134.5
(OCH2CH=CH2), 128.4, 127.8, 127.1 (C Ar), 117.3 (OCH2CH=CH2), 96.6 (C-1), 95.0 (C-2),
90
69.7 (C-4), 69.2 (CH2Ar), 68.9 (OCH2CH=CH2), 65.4 (C-5), 38.0, 28.0 (CH3COCH2CH2CO),
29.7 (CH3COCH2CH2CO), 17.8 (C-6). HRMS gave unexpected results which we are currently
investigating.
Allyl 3-O-benzyl-4-O-levulinoyl-α-L-glucopyranoside (40)
Triflate 39 (50 mg, 0.097 mmol) was dissolved in anhydrous DMF (2 mL). Tetrabutylammonium
nitrite (223 mg, 8 equiv) was added and the reaction flask was sealed and stirred at 50 oC for 6
hr. The reaction was then diluted with EtOAc (10 mL) and washed with water (4 × 2 mL). The
organic phase was dried over Na2SO4 and concentrated. The crude was purified by silica
chromatography (2:8 EtOAc:Hex) to give the eliminated product 41 (4.35 mg, 12%) and 40 (1.5
mg, 4%) both as oil. [α]D 39.0 (c 1.0, MeOH) 1H NMR for 41 (CDCl3, 400 MHz, 295 K) δH
7.33-7.25 (m, 5H, Ar), 5.96-5.86 (m, 1H, OCH2CH=CH2), 5.33-5.20 (m, 2H, OCH2CH=CH2),
4.86 (d, 1H, J = 3.3 Hz, H-1), 4.81 (d, 1H, J = 11.7 Hz, CH2Ar), 4.75 (t, 1H, J = 8.6 Hz, H-4)
4.68 (d, 1H, J = 11.7 Hz, CH2Ar), 4.19 (bdd, 1H, J = 5.3, 12.7 Hz, OCHHCH=CH2), 4.03 (bdd,
1H, = 6.3, 12.7 Hz, OCHHCH=CH2), 4.81-3.77 (m, 1H, H-5), 3.71-3.66 (m, 2H, H-2, H-3),
2.73-2.31 (m, 4H, CH3COCH2CH2CO), 2.13 (s, 3H, CH3COCH2CH2CO), 1.13 (d, 3H, J = 6.3
Hz, H-6). 13C NMR (CDCl3, 100 MHz, 295 K): δC 206.4 (CH3COCH2CH2CO), 172.8
(CH3COCH2CH2CO), 138.5 (4o Ar), 133.5 (OCH2CH=CH2),128.5-127.6 (C Ar), 118.1
(OCH2CH=CH2), 97.2 (C-1), 80.3 (C-3), 75.1 (C-4), 74.5 (CH2Ar), 72.8 (C-2), 68.6
91
(OCH2CH=CH2), 65.9 (C-5), 37.8, 27.9 (CH3COCH2CH2CO), 29.8 (CH3COCH2CH2CO), 17.2
(C-6). HRMS gave unexpected results which we are currently investigating.
Allyl 3-O-benzoyl-4-O-levulinoyl-α-L-rhamnopyranoside (37)
Diol 37 (1.02 g , 3.37 mmol) was dissolved in anhydrous DCM (10 mL) and pyridine (2 mL, 7.3
equiv) and placed under N2. BzCl (500 μL, 1.3 equiv) was added and the reaction was allowed to
stir overnight at room temperature. The next morning another addition of BzCl (500 μL, 1.3
equiv) was added and stirred for 6 hours. The reaction was diluted in DCM (10 mL) and was
washed with 1N HCl (30 mL) followed by sat’d NaHCO3 (50 mL). The organic phase was dried
and concentrated and was purified by silica chromatography (1:9 to 2:7 EtOAc:Hexanes) to give
benzoate 37 (0.87 g 64%) as a yellow oil. [α]D -11.7 (c 1.0, MeOH) 1H NMR for 37 (CDCl3, 400
MHz, 295 K) δH 8.00 (d, 2H, J = 7.4 Hz, Bz), 7.53 (t, 1H, J = 6.1 Hz, OBz), 7.42 (t, 2H, J = 8.0
Hz, OBz), 5.96-5.85 (1H, m, OCH2CH=CH2), 5.43 (dd, 1H, J = 3.2, 10.1 Hz), 5.34-5.21 (m, 2H,
OCH2CH=CHH), 4.86 (s, 1H, H-1), 4.26-4.18 (m, 2H, OCHHCH=CH2, H-2), 4.04 (dd, 1H, J =
6.1, 12.9 Hz, OCHHCH=CH2), 3.99-3.91 (m, 1H, H-5), 2.67-2.31 (m, 4H, CH3COCH2CH2CO),
2.04 (s, 3H, CH3COCH2CH2CO), 1.25 (d, 3H, J = 6.3 Hz). 13C NMR (CDCl3, 100 MHz, 295 K):
δC 206.1 (CH3COCH2CH2CO), 172.0 (CH3COCH2CH2CO), 165.5 (Bz), 133.4
92
(OCH2CH=CH2), 133.4, 129.8, 128.5 (Ar), 129.4 (4o Ar), 117.7 (OCH2CH=CH2), 98.4 (C-1),
72.5 (C-3), 71.2 (C-4), 69.6 (C-2), 68.2 (OCH2CH=CH2), 66.4 (C-5), 37.8, 27.9
(CH3COCH2CH2CO), 29.6 (CH3COCH2CH2CO), 17.3 (C-6). HRESIMS calculated for C21H26O8
[M+Na]+ 429.1519, found 429.1503.
Allyl 3-O-benzoyl-4-O-levulinoyl-2-O-trifyl-α-L-rhamnopyranoside (43)
Alcohol 42 (83 mg, 0.2 mmol) was dissolved in anhydrous DCM (2 mL) and anhydrous pyridine
(100 μL, 6.2 equiv). The reaction was sealed under N2 and cooled to -20 oC. Tf2O (70 μL, 2
equiv) was added drop wise to the reaction. The reaction was allowed to warm to 0 oC over 45
min. The reaction was diluted in DCM (5 mL) and was washed with 1N HCl (5 mL) followed by
sat’d NaHCO3 (5 mL). The organic phase was dried and concentrated to give triflate 43 (103 mg,
92%) as a clear oil. (c 1.0, MeOH) 1H NMR for 43 (CDCl3, 400 MHz, 295 K) 8.00 (d, 2H, J =
7.1 Hz, OBz), 7.57 (t, 1H, J = 6.2 Hz, OBz), 7.43 (t, 2H, J = 8.0 Hz, OBz), 5.94-5.84 (m, 1H,
OCH2CH=CH2), 5.60 (dd, 1H, J = 3.1, 10.2 Hz, H-3), 5.36-5.26 (m, 2H, OCH2CH=CH2), 5.24
(t, 1H, J = 9.9 Hz, H-4), 5.17 (bs, 1H, H-2), 5.00 (d, 1H, J = 1.4 Hz, H-1), 4.24 (bdd, 1H, J = 5.2,
12.7 Hz, OCHHCH=CH2), 4.07 (bdd, 1H, J = 6.3, 12.7 Hz, OCHHCH=CH2), 4.03-3.96 (m, 1H,
H-5), 2.70-2.45 (m, 4H, CH3COCH2CH2CO), 2.03 (s, 3H, CH3COCH2CH2CO), 1.28 (d, 3H, J =
6.2 Hz, H-6). 13C NMR (CDCl3, 100 MHz, 295 K): δC 206.1 (CH3COCH2CH2CO), 171.6
(CH3COCH2CH2CO), 165.5 (Bz), 133.7, 129.9, 128.5 (Ar), 132.5 (OCH2CH=CH2), 128.6 (4o
93
Ar), 118.8 (OCH2CH=CH2), 95.6 (C-1), 82.5 (C-2), 70.2 (C-4), 68.8 (OCH2CH=CH2), 68.7 (C3), 66.8 (C-5), 37.8, 27.8 (COCH2CH2CO), 29.5 (CH3COCH2CH2CO), 17.1 (C-6).
Allyl 3-O-benzoyl-4-O-levulinoyl-2-O-trifyl-α-L-glucoopyranoside (46), Allyl 2-6dianhydro-3-O-benzoyl-4-O-levulinoyl-α-L-erythro-hex-3-enopyranoside (47)
Triflate 43 (53 mg, 0.098 mmol) was dissolved in anhydrous DMF (2 mL). Tetrabutylammonium
nitrite (743 mg, 25 equiv) was added and the reaction flask was sealed and stirred at rt for 24 hr.
The reaction was then diluted with EtOAc (10 mL) and washed with water (4 × 2 mL). The
organic phase was dried over Na2SO4 and concentrated. The crude was purified by silica
chromatography (3:7 EtOAc:Hex) to give inverted sugar 46 (9.9 mg, 23%) and the eliminated
product 47 (10.6 mg, 28%) both as oils. [α]D -39.0 (c 1.0, MeOH) 1H NMR for 46 (CDCl3, 400
MHz, 295 K) δH 8.00 (bd, 2H, J = 8.3 Hz, Bz), 7.60 (bt, 1H, J = 8.3 Hz, Bz), 7.48 (t, 1H, J = 8.3
Hz, Bz), 5.97-5.86 (m, 1H, OCH2CH=CH2), 5.45 (t, 1H, J = 9.7 Hz, H-3), 5.35-5.31 (m, 2H,
OCH2CH=CH2), 4.93 (t, 1H, J = 9.3, H-4), 4.92 (d, 1H, J = 2.8 Hz, H-1), 4.27-4.22 (m, 1H,
OCHHCH=CH2), 4.08-4.03 (m, 1H, OCHHCH=CH2), 3.98-3.90 (m, 1H, H-5), 3.79 (td, 1H, J =
94
3.5, 9.6 Hz, H-2) 2.60-2.31 (m, 4H, CH3COCH2CH2CO), 2.21 (d, 1H, J = 11.1 Hz, OH-2), 2.03
CH3COCH2CH2CO), 1.22 (d, 3H, J = 6.3 Hz, H-6). 13C NMR (CDCl3, 100 MHz, 295 K): δC
206.0 (CH3COCH2CH2CO), 171.8 (CH3COCH2CH2CO), 166.8 (Bz), 133.3 (OCH2CH=CH2),
133.2, 129.9, 128.4 (Ar), 129.5 (4o Ar), 118.2 (OCH2CH=CH2), 97.4 (C-1), 74.0 (C-3), 73.2 (C4), 71.5 (C-2), 68.8 (OCH2CH=CH2), 65.8 (C-5), 37.8, 27.9 (CH3COCH2CH2CO), 29.5
(CH3COCH2CH2CO), 17.1 (C-6). HRESIMS calculated for C21H26O8 [M-C7H6O2+H]+ 285.1340,
found 285.1323.
[α]D -54.2 (c 0.9, MeOH) 1H NMR for 47 (CDCl3, 600 MHz, 295 K) δH 8.03 (bd, 2H, J = 6.8
Hz, Bz), 7.62-7.40 (m, 3H, Bz), 6.01-5.93 (m, 1H, OCH2CH=CH2), 5.90 (bs, 1H, H-2) 5.53 (d,
1H, J = 9.0, H-4), 5.37-5.23 (m, 2H, OCH2CH=CH2) 5.28 (bs, 1H, H-1), 4.32-4.22 (m, 2H,
OCHHCH=CH2, H-5), 4.15-4.10 (m, 1H, OCHHCH=CH2), 2.69-2.42 (m, 4H,
CH3COCH2CH2CO), 2.09 (s, 3H, CH3COCH2CH2CO), 1.30 (d, 3H, J = 5.0 Hz, H-6). 13C NMR
(CDCl3, 100 MHz, 295 K): δC 206.0 (CH3COCH2CH2CO), 172.4 (CH3COCH2CH2CO), 163.7
(C=O), 147.3 (C-3), 134.5 (OCH2CH=CH2), 128.4, 127.8, 127.1 (C Ar), 128.9 (4o Ar), 117.3
(OCH2CH=CH2), 115.0 (C-2), 93.9 (C-1), 69.4 (C-4), 69.2 (OCH2CH=CH2), 66.1 (C-5), 37.7,
27.8 (CH3COCH2CH2CO), 29.7 (CH3COCH2CH2CO), 17.8 (C-6). HRESIMS calculated for
C21H24O7 [M+Na]+ 411.1414, found 411.1398.
95
Allyl 3-O-benzoyl-4-O-levulinoyl-2-O-mesyl-α-L-rhamnopyranoside (44)
The alcohol 42 (120 mg, 0.34 mmol) was dissolved in anhydrous DCM (2 mL) and anhydrous
pyridine (150 μL, 5.4 equiv). The reaction was sealed under N2 and cooled to 0 oC. MsCl (52 μL,
2 equiv) was added drop wise to the reaction. The reaction was allowed to warm to RT and
stirred for 3 hr. The reaction was diluted in DCM (5 mL) and was washed with 1N HCl (5 mL)
followed by sat’d NaHCO3 (5 mL). The organic phase was dried and concentrated to give the
mesylate 44 (148 mg, 90%) as a slightly orange powder. [α]D -4.1 (c 1.0, MeOH) 1H NMR for 44
(CDCl3, 400 MHz, 295 K) δH 8.00 (bd, 2H, J = 6.0 Hz, Bz), 7.58 (bt, 1H, J = 7.3 Hz, Bz), 7.46
(t, 1H, J = 7.9 Hz, Bz), 5.97-5.86 (m, 1H, OCH2CH=CH2), 5.55 (dd, 1H, J = 3.3, 10.3 Hz, H-3),
5.34-5.20 (m, 2H, OCH2CH=CH2), 5.27 (t, 1H, J = 10.0 Hz, H-4), 5.13 (bs, 1H, H-2), 5.01 (bs,
1H, H-1), 4.23 (bdd, 1H, J = 5.5, 12.7 Hz, OCHHCH=CH2), 4.07 (bdd, 1H, J = 6.5, 12.7 Hz,
OCHHCH=CH2), 2.92 (s, 3H, SO2CH3), 2.61-2.33 (m, 4H, CH3COCH2CH2CO), 2.05 (s, 3H,
CH3COCH2CH2CO), 1.25 (d, 3H, J = 6.3 Hz, H-6). 13C NMR (CDCl3, 100 MHz, 295 K): δC
205.9 (CH3COCH2CH2CO), 171.8 (CH3COCH2CH2CO), 165.5 (Bz), 133.6 (OCH2CH=CH2),
132.9, 129.8, 128.6 (Ar), 129.0 (4o Ar), 118.3 (OCH2CH=CH2), 96.7 (C-1), 76.1 (C-2), 70.6 (C4), 69.6 (C-3), 68.8 (OCH2CH=CH2), 66.6 (C-5), 38.4 (SO2CH3), 38.4, 27.8
(CH3COCH2CH2CO), 29.6 (CH3COCH2CH2CO), 17.2 (C-6). HRESIMS calculated for
C22H28O10S [M+Na]+ 507.1295, found 507.1281.
96
3-O-Benzoyl-4-O-levulinoyl-2-O-mesyl-α-L-rhamnopyranoside (48)
Sugar 48 (101 mg, 0.206 mmol) was dissolved in 10% aqueous ethanol with DABCO (23 mg,
0.2 equiv) and RhCl(PPh3)3 (13 mg, 0.07 equiv) and refluxed under nitrogen for 4 hr. The
reaction was allowed to cooled and then concentrated. The reaction was diluted with EtOAc (5
mL) and filtered through Celite with copious washes of EtOAc. The organic phase was washed
with 1 M HCl (5 mL) and then dried over Na2SO4 and concentrated to an red oil. The oil was
dissolved in acetone and water (10:1) and yellow mercuric oxide (55 mg, 1.25 equiv) and
mercury chloride (55 mg, 1 equiv) and was stirred at room temperature for 2 hr. The reaction
was filtered through Celite with copious washes of acetone. The filtrate was concentrated and
then diluted with EtOAc (10 mL) and washed with water (3 × 10 mL). The organic phase was
dried over Na2SO4 and concentrated to an oil. The product was purified by silica
chromatography (2:8 EtOAc:Hex) to give the hemiacetal 48 (71%, 65 mg) as a white powder. 1H
NMR for 48 (CDCl3, 400 MHz, 295 K) δH 7.98 (bd, 2H, J = 6.0 Hz, Bz), 7.55 (bt, 1H, J = 7.3
Hz, Bz), 7.42 (t, 1H, J = 7.9 Hz, Bz), 5.56 (dd, 1H, J = 3.1 Hz, H-3), 5.37 (bs, 1H, H-1), 5.23 (t,
1H, J = 10.0 Hz, H-4), 5.11 (dd, 1H, J = 1.9, 3.2 Hz, H-2), 4.21-4.17 (m, 1H, H-5), 3.58 (d, 1H, J
= 4.1 Hz, OH-1), 2.94 (s, 3H, SO2CH3), 2.65-2.33 (m, 4H, CH3COCH2CH2CO), 2.04 (s, 3H,
CH3COCH2CH2CO), 1.25 (d, 3H, J = 6.3 Hz, H-6). 13C NMR (CDCl3, 100 MHz, 295 K): δC
205.9 (CH3COCH2CH2CO), 171.8 (CH3COCH2CH2CO), 165.5 (Bz), 133.6 (OCH2CH=CH2),
132.9, 129.8, 128.6 (Ar), 129.0 (4o Ar), 118.3 (OCH2CH=CH2), 96.7 (C-1), 76.1 (C-2), 70.6 (C-
97
4), 69.6 (C-3), 68.8 (OCH2CH=CH2), 66.6 (C-5), 38.4 (SO2CH3), 38.4, 27.8
(CH3COCH2CH2CO), 29.6 (CH3COCH2CH2CO), 17.2 (C-6). HRESIMS calculated for
C19H24O10S [M+Na]+ 467.0993, found 467.0979.
3-O-Benzoyl-4-O-levulinoyl-2-O-mesyl-1-O-trichloroacetimidate-α-L-rhamnopyranoside
(50)
Hemiacetal 48 (52 mg, 0.11 mmol) was dissolved in anhydrous DCM (2 mL) and was placed
under N2. Cl3CCN (35 μL, 3.25 equiv) and 1,8-diazabicyclo(5.4.0)undec-7-ene (5 μL, 0.25
equiv) and was stirred overnight. The reaction was concentrated and then purified with silica
chromatography (1:9 EtOAc:Hex w/ 0.1% Et3N) to give donor 50 (46 mg, 77%) as an off white
powder. 1H NMR for 50 (CDCl3, 400 MHz, 295 K) δH 8.79 (s, 1H, NH) 8.00 (bd, 2H, J = 8.5
Hz, Bz), 7.57 (tt, 1H, J = 1.3, 7.4 Hz, Bz), 7.44 (t, 1H, J = 7.5 Hz, Bz), 6.38 (d, 1H, J = 1.7 Hz,
H-1), 5.54 (dd, 1H, J = 3.4, 10.7 Hz, H-3), 5.35 (t, 1H, J = 10.1 Hz, H-4), 5.31 (dd, 1H, J = 1.9,
3.2 Hz, H-2), 4.22-4.15 (m, 1H, H-5), 3.02 (s, 3H, SO2CH3), 2.73-2.35 (m, 4H,
CH3COCH2CH2CO), 2.04 (s, 3H, CH3COCH2CH2CO), 1.33 (d, 3H, J = 6.3 Hz, H-6).13C NMR
(CDCl3, 100 MHz, 295 K): δC 205.9 (CH3COCH2CH2CO), 171.8 (CH3COCH2CH2CO), 165.6
98
(Bz), 159.8 (C=NH), 132.9, 129.8, 128.6 (Ar), 129.0 (4o Ar), 94.8 (C-1), 74.0 (C-2), 69.7 (C-4),
69.6 (C-5), 69.2 (C-3), 38.6 (SO2CH3), 37.8, 27.8 (CH3COCH2CH2CO), 29.6
(CH3COCH2CH2CO), 17.2 (C-6).
Methyl 3-O-(3-O-benzoyl-4-O-levulinoyl-2-O-mesyl-α-L-rhamnopyranoside)- 2-O-benzoyl4,6-di-O-benzyl-α-D-mannopyranoside (53)
Acceptor 3 (12 mg, 0.02 mmol), and donor 5 (32 mg, 0.05 mmol), was dissolved in anhydrous
DCM (500 μL) and stirred under N2 for 15 min at RT. A 0.01 M solution of TMSOTf was
created and added dropwise (490 μL, 0.12 equiv) to the reaction. The reaction was stirred for 3
hr and then quenched with Et3N (5 μL) and stored in the freezer overnight. The next morning the
reaction was concentrated purified by silica chromatography (2:8 EtOAc:Hex), however,
purification was unsuccessful, giving disaccharide 53 (7.7 mg, 33%, estimated by NMR) with 3
as an oil. 1H NMR for 53 (CDCl3, 400 MHz, 295 K) δH 8.04 (dd, 2H, J = 1.3, 8.4 Hz, Bz), 7.97
(dd, 3H, J = 1.3, 7.2 Hz, Bz), 7.59-7.49 (m, 3H, Bz), 7.45-7.31 (m, 13H, Bz, Bn), 5.54 (bt, 1H, J
= 2.9 Hz, H-2), 5.45 (dd, 1H, J = 3.6, 10.6 Hz, H-3’), 5.35 (d, 1H, J = 1.6 Hz, H-1’), 5.18 (t, 1H,
J = 10.2 Hz, H-4’), 4.87 (d, 1H, J = 1.9 Hz, H-1), 4.83-4.82 (m, 2H, H-2’, CHHAr), 4.79 (d, 1H,
J = 11.7 Hz, CHHAr), 4.60-4.57 (m, 2H, 2 × CHHAr), 4.34 (dd, 1H, J = 2.8, 9.4 Hz, H-3), 4.14-
99
4.02 (m, 2H, H-4, H-5’), 3.95 (dd, 1H, J = 3.7, 10.7 Hz, H-6A), 3.90-3.87 (m, 1H, H-5), 3.81
(dd, 1H, J = 1.7, 10.7 Hz, H-6B), 3.40 (s, 3H, OMe), 3.17 (s, 3H, OMs), 2.55-2.35 (m, 4H,
CH3COCH2CH2CO), 1.98 (s, 3H, CH3COCH2CH2CO), 1.10 (d, 3H, J = 6.2 Hz, H-6’). 13C NMR
(CDCl3, 100 MHz, 295 K): δC 205.9 (CH3COCH2CH2CO), 171.5 (CH3COCH2CH2CO), 165.1
(Bz), 165.5 (Bz), 138.1, 137.6, 129.1, 129.0 (4o Ar), 130.0, 129.8, 128.6-127.5 (Ar), 98.9 (C-1, J
= 174.3 Hz), 93.6 (C-1’, J = 175.3 Hz), 77.5 (C-2’), 75.6 (CH2Ar), 73.5 (CH2Ar), 73.4 (C-3),
73.7 (C-4), 71.6 (C-5), 70.6 (C-4’), 68.9 (C-3’), 68.7 (C-6), 67.6 (C-2), 66.8 (C-5’), 55.1 (OMe),
38.6 (SO2CH3), 37.8, 27.8 (CH3COCH2CH2CO), 29.6 (CH3COCH2CH2CO), 17.1 (C-6’).
HRESIMS calculated for C47H52O16S [M+Na]+ 927.2868, found 927.2835.
.
Allyl 3-O-benzoyl-4-O-levulinoyl-2-O-tosyl-α-L-rhamnopyranoside (45)
Alcohol 42 (201 mg, 0.49 mmol) was dissolved in anhydrous DCM (2 mL) and anhydrous
pyridine (200 μL, 5 equiv). TsCl (52 μL, 2 equiv) was added heated to 30 oC. The reaction was
stirred overnight then cooled to RT. The reaction was diluted in DCM (5 mL) and was washed
with 1N HCl (5 mL) followed by sat’d NaHCO3 (5 mL). The organic phase was dried and
concentrated. The crude was purified by silica chromatography (2:8 EtOAc:Hex) to give tosylate
45 (238 mg, 87%) as slight orange powder. [α]D -14.2 (c 1.0, MeOH) 1H NMR for 45 (CDCl3,
100
400 MHz, 295 K) δH 7.72 (dd, 2H, J = 1.2, 8.1 Hz, Bz), 7.60 (d, 2H, J = 8.3 Hz, Ts), 7.54 (dt,
1H, J = 1.3, 7.5 Hz, Bz), 7.38 (t, 2H, J = 7.8 Hz, Bz), 6.86 (d, 2H, J = 8.2 Hz, Ts), 5.92-5.82 (m,
1H, OCH2CH=CH2), 5.32-5.19 (m, 4H, H-3, H-4, OCH2CH=CH2), 5.01 (d, 1H, J = 1.4 Hz, H1), 4.84 (dd, 1H, J = 1.8, 3.3 Hz H-2), 4.19 (bdd, 1H, J = 5.5, 12.7 Hz, OCHHCH=CH2), 4.05
(bdd, 1H, J = 6.5, 12.7 Hz, OCHHCH=CH2), 3.93-3.86 (m, 1H, H-5), 2.66-2.26 (m, 4H,
CH3COCH2CH2CO), 2.08 (s, 3H, OTs), 2.01 (s, 3H, CH3COCH2CH2CO), 1.25 (d, 3H, J = 6.3
Hz, H-6). 13C NMR (CDCl3, 100 MHz, 295 K): δC 205.9 (CH3COCH2CH2CO), 171.8
(CH3COCH2CH2CO), 165.5 (Bz), 144.7, 132.9, 129.8, 128.6 (4o Carmo), 133.6 (OCH2CH=CH2),
129.0 (Ar), 118.0 (OCH2CH=CH2), 96.8 (C-1), 76.2 (C-2), 70.4 (C-4), 69.6 (C-3), 68.5
(OCH2CH=CH2), 66.4 (C-5), 37.8, 27.8 (CH3COCH2CH2CO), 29.5 (CH3COCH2CH2CO), 17.2
(C-6). HRESIMS calculated for C28H32O10S [M+Na]+ 583.1608, found 583.1608
3-O-Benzoyl-4-O-levulinoyl-2-O-tosyl-α-L-rhamnopyranoside (49)
Sugar 45 (223 mg, 0.39 mmol) was dissolved in 10% aqueous ethanol with DABCO (44 mg, 0.2
equiv) and RhCl(PPh3)3 (24 mg, 0.07 equiv) and refluxed under nitrogen for 4 hr. The reaction
was allowed to cooled and then concentrated. The reaction was diluted with EtOAc (5 mL) and
filtered through Celite with copious washes. The organic phase was washed with 1 M HCl (5
101
mL) and then dried over Na2SO4 and concentrated to an red oil. The oil was dissolved in acetone
and water (10:1) and yellow mercuric oxide (104 mg, 1.25 equiv) and mercury chloride (104 mg,
1 equiv) and was stirred at room temperature for 2 hr. The reaction was filtered through Celite
with copious washes of acetone. The filtrate was concentrated and then diluted with EtOAc (10
mL) and washed with water (3 × 10 mL). The organic phase was dried over Na2SO4 and
concentrated to an oil. The product was purified by silica chromatography (2:8 EtOAc:Hex) to
give henmiacetal 49 (152 mg, 75%) as a white powder. 1H NMR for 49 (CDCl3, 400 MHz, 295
K) 7.69 (dd, 2H, J = 1.0, 8.4 Hz, Bz), 7.58 (d, 2H, J = 8.4 Hz, Ts), 7.53 (tt, 2H, J = 1.2, 7.4 Hz,
Bz), 7.36 (bt, 2H, J = 7.7 Hz, Bz), 6.83 (d, 2H, J = 7.7 Hz, Ts), 5.37 (bs, 1H, H-1), 5.35 (dd, 1H,
J = 3.5 Hz, 10.4 Hz), 5.21 (t, 1H, J = 9.9 Hz, H-4), 4.85 (dd, 1H, J = 1.9, 3.3 Hz, H-2), 4.14-4.10
(m, 1H, H-5), 2.61-2.27 (m, 4H, CH3COCH2CH2CO), 2.04 (s, 3H, Ts), 1.93
(CH3COCH2CH2CO), 1.21 (d, 3H, J = 6.1 Hz, H-6’).13C NMR (CDCl3, 100 MHz, 295 K): δC
206.4 (CH3COCH2CH2CO), 171.8 (CH3COCH2CH2CO), 165.2 (Lev), 165.0 (Bz), 144.6, 132.5,
128.8 (4o Carmo), 133.3, 129.8, 129.7, 128.2, 127.2 (Carmo), 92.4 (C-1), 76.7 (C-2), 70.5 (C-4),
69.3 (C-3), 66.2 (C-5), 37.8, 27.8 (CH3COCH2CH2CO), 29.5 (CH3COCH2CH2CO), 21.5 (Ts),
17.1 (C-6). HRESIMS calculated for C25H28O10S [M+Na]+ 543.1295, found 543.1299.
102
3-O-Benzoyl-4-O-levulinoyl-2-O-tosyl-1-O-trichloroacetimidate-α-L-rhamnopyranoside
(51)
Hemiacetal 49 (121 mg, 0.23 mmol) was dissolved in anhydrous DCM (2 mL) and was placed
under N2. Cl3CCN (74 μL, 3.25 equiv) and 1,8-diazabicyclo(5.4.0)undec-7-ene (11 μL, 0.25
equiv) and was stirred overnight. The reaction was concentrated and then purified with silica
chromatography (1:9 EtOAc:Hex w/ 0.1% Et3N) to give donor 51 (117 mg, 78%) as an off white
powder. 1H NMR for 51 (CDCl3, 400 MHz, 295 K) δH 8.77 (s, 1H, NH), 7.76 (dd, 2H, J = 1.3,
8.4 Hz, Bz), 7.64 (d, 2H, J = 8.2 Hz, Ts), 7.57 (tt, 1H, J = 1.3, 7.4 Hz, Bz), 7.39 (bt, 2H, J = 7.9
Hz, Bz), 6.91 (d, 2H, J = 8.1 Hz, Ts), 6.39 (d, 1H, J = 1.4 Hz, H-1), 5.38-5.34 (m, 2H, H-3, H-4)
5.07 (t, 1H, J = 2.5 Hz, H-2), 4.15-4.10 (m, 1H, H-5), 2.71-2.29 (m, 4H, CH3COCH2CH2CO),
2.14 (s, 3H, Ts), 2.02 (CH3COCH2CH2CO), 1.30 (d, 3H, J = 6.2 Hz, H-6’). 13C NMR (CDCl3,
100 MHz, 295 K): δC 205.8 (CH3COCH2CH2CO), 171.6 (CH3COCH2CH2CO), 165.2 (Lev),
165.0 (Bz), 159.6 (C=NH), 145.0, 132.3, 128.7 (4o Carmo), 133.4, 129.9, 129.8, 128.3-127.9
(Carmo), 94.9 (C-1), 74.3 (C-2), 69.6 (C-4), 69.3 (C-3, C-5), 37.8, 27.8 (CH3COCH2CH2CO), 29.5
(CH3COCH2CH2CO), 21.5 (Ts), 17.3 (C-6).
103
Methyl 3-O-(3-O-benzoyl-4-O-levulinoyl-2-O-tosyl-α-L-rhamnopyranoside)-2-O-benzoyl4,6-di-O-benzyl-α-D-mannopyranoside (54)
Acceptor 3 (30 mg, 0.06 mmol), and donor 5 (55 mg, 0.08 mmol), was dissolved in anhydrous
DCM (1 mL) and stirred under N2 for 15 min at RT. A 0.01 M solution of TMSOTf was created
and added dropwise (970 μL, 0.12 equiv) to the reaction. The reaction was stirred for 3 hr and
then quenched with Et3N (5 μL) and stored in the freezer overnight. The next morning the
reaction was concentrated purified by silica chromatography (2:8 EtOAc:Hex) to give
disaccharide 53 (43 mg, 70%) as an oil. [α]D -16.5 (c 1.0, MeOH) 1H NMR for 54 (CDCl3, 400
MHz, 295 K) δH 8.11 (bd, 2H, J = 7.2 Hz, Bz), 7.76 (bd, 2H, J = 7.2 Hz), 7.62-7.52 (m, 5H, Ts,
Bz), 7.44-7.19 (m, 15H, Ts × 3, Bz), 6.82 (d, 2H, J = 8.3 Hz, Ts), 5.45 (bt, 1H, J = 3.5 Hz, H-2),
5.32 (dd, 1H, J = 3.2, 10.2 Hz, H-3’), 5.21 (bs, 1H, 1-H’), 5.21 (t, 1H, J = 10.1 Hz), 4.88 (bd, 1H
J = 1.7 Hz, H-1), 4.80 (d, 1H, J = 10.9 Hz, CHHAr), 4.77 (d, 1H, J = 12.0 Hz, CHHAr), 4.66
(dd, 1H, J = 1.8, 3.3 Hz, H-2’), 4.60 (d, 1H, J = 11.4 Hz, CHHAr), 4.57 (d, 1H, J = 12.2 Hz,
CHHAr), 4.31 (dd, 1H, J = 3.2, 9.8 Hz, H-3), 4.12-4.01 (m, 2H, H-4, H-5’), 3.94 (dd, 1H, J =3.8,
10.8 Hz, H-6A), 3.87 (m, 1H, H-5), 3.80 (bd, 1H, J = 10.8 Hz, H-6B), 3.38 (s, 3H, OMe), 2.552.17 (m, 4H, CH3COCH2CH2CO), 2.14 (s, 3H, Ts), 2.02 (CH3COCH2CH2CO), 1.07 (d, 3H, J =
6.2 Hz, H-6’). 13C NMR (CDCl3, 100 MHz, 295 K): δC 205.9 (CH3COCH2CH2CO), 171.8
(CH3COCH2CH2CO), 165.6 (Bz), 165.0 (Bz), 144.6, 138.1, 137.7, 132.5, 129.6, 129.0 (4o Carmo),
104
130.1, 129.8, 129.7, 129.0-127.6 (Carmo), 98.7 (C-1, J = 174.3 Hz), 93.6 (C-1’, J = 173.9 Hz),
76.0 (C-2’), 75.4 (CH2Ar), 73.6 (C-4), 73.5 (C-3), 73.5 (CH2Ar), 71.5 (C-5), 70.4 (C-4’), 69.3
(C-3’), 68.6 (CH2Ar), 67.9 (C-2’), 66.5 (C-5’), 60.4 (C-6), 55.1 (OMe), 37.7, 27.6
(CH3COCH2CH2CO), 29.5 (CH3COCH2CH2CO), 21.6 (Ts), 17.1 (C-6’). HRESIMS calculated
for C53H56O16S [M+Na]+ 1003.3181, found 1003.3163.
105
Chapter 6- References
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