GLYCOSYLATION METHODS
IN
OLIGOSACCHARIDE SYNTHESIS
by Inmaculada Robina
Department of Organic Chemistry. University of Seville
-2-
GLYCOSYLATION METHODS IN OLIGOSACCHARIDE SYNTHESIS
Introduction
Glycoconjugates are biopolymers formed by an oligosaccharide moiety joined to a protein
(glycoproteins) or to a lipid moiety (glycolipids). These biopolymers together with proteins and
nucleic acids are mainly responsible of information transfer between cells, which is a
fundamental process of life and central to all cellular systems.
Nowadays it is well known that complex oligosaccharides in the form of glycolipids and
glycoproteins are present in the membranes of cells and can mediate a large number of diverse
and important biological functions. Oligosaccharides play a major role in inflammation, immune
response, metastasis, fertilization and many other important biomedical processes. Specific
carbohydrates cover different kinds of functions. For instance, they act as markers of certain
types of tumours, other act as signal molecules of symbiotic processes such as the symbiosis
between Rhizobium bacteria and legume plants; others are binding site for bacterial and viral
pathogens, etc…
The area of organic chemistry that deals with the study, preparation and biological role of
sugars, from monosaccharides to complex oligosaccharides and their analogues, is called
Glycobiology.
The important role of carbohydrates in Biology and Biomedicine has been a major incentive
for devising new methods for the chemical and enzymatic synthesis of this class of molecules.
The biological role of sugars depends on many factors. Compared with other biopolymers
such as nucleic acids, proteins and peptides, in which their biological activity depends on their
sequence of nucleotides or amino acids, in the case of oligosaccharides, the situation is more
complex. For oligosaccharides, besides the sequence of the monomeric structures, other aspects
such as the functional groups and their stereochemistry, the conformation of the sugars
ramification, the stereoselective formation of glycosidic linkages, etc… must be considered.
All these facts have made the area of oligosaccharide synthesis an ideal and challenging area
for the development and testing new synthetic methodologies.
-3-
This course is divided in three lessons:
1.
General Aspects of Oligosaccharide Synthesis
2.
Different Procedures of Glycosylation Reactions by Direct Activation
3.
Synthetic Strategies for the Assembly of Oligosaccharides
Bibliography (Books)
1.- Preparative Carbohydrate Chemistry, Ed. Stephen Hanessian. University of
Montreal, Canada. Marcel Dekker, Inc. New York, 1997
2.- Carbohydrate Chemistry, Ed. G. –J. Boons, Blackie Academic Professional,
1998
3.- Modern Methods in Carbohydrate Syntheses, Eds. S. H. Khan and R. A.
O`Neill. Haword Academic Press, 1996
-4-
Lesson 1. General aspects of oligosaccharide synthesis
1.
2.
3.
4.
Formation of a glycosidic bond
General mechanistic pathway for glycosidic bond formation
Choices, challenges and problems of the glycosidic bond
Structure and reactivity of glycosyl donors and of glycosyl acceptors used in
oligosaccharide synthesis
5. Promoters, solvents and experimental conditions
6. Anomeric control in chemical glycosylations. Methods for stereoselective formation of
glycosidic linkages.
6.1. Preparation of 1,2-trans-glycosides by neighbouring group participation
6.2. In situ anomerization for the synthesis of α-glycosides (Lemieux)
6.3. Heterogeneous catalysis (Paulsen).
6.4. Stereoselective preparation of α- and β-glycosides by participation of the solvent
6.5. Intramolecular aglycone delivery approach
7. Common protecting groups used in oligosaccharide synthesis
1. Formation of a glycosidic bond
This bond is formed by a nucleophilic displacement of a leaving group (X) attached to the
anomeric carbon of a sugar moiety by an alcohol ROH, or by the OH group of a partially
protected sugar moiety. The compound that “gives” the glycosyl moiety, is called the glycosyl
donor, and the alcohol that receives it, is known as glycosyl acceptor. The reaction generally is
performed in the presence of an activator called “promoter”. The role of the promoter is to assist
the departure of the leaving group. Promoters are often used in catalytic amounts, although in
some instances they are used stoichiometrically. In some cases, other additives such as
molecular sieves or any base that may act as acid scavenger are used.
There are many methods available for glycosidic bond formation. In this course, we will
discuss the most important and the widely applicable ones.
O
G
X +
HO-R'
OR
O
G
OR
glycosyl donor
(electrophile)
X +
O
HO
promoter
solvent
OR'
promoter
solvent
O
G
OR'
R
O
O
G
R
O
OR'
glycosyl acceptor
(nucleophile)
Scheme 1
The synthesis of disaccharides and oligosaccharides in general, involves the linking of two
polyfunctional compounds. It is much more complicated than the synthesis of other biopolymers
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such as peptides or nucleic acids because of the greater number of possibilities for the
combination of monomeric units and because the glycosidic linkages have to be introduced in a
stereospecific way.
2. General mechanistic pathway for glycosidic bond formation1
The General Mechanistic Pathways for Glycosidic Bond Formation is represented in Scheme
2. Over 90% of all the glycosylations reported, formally proceed via this general mechanistic
pathway. There are some exceptions such as in situ anomerization, intramolecular aglycon
delivery and the use of additives such as acetonitrile, which appears to react at the anomeric
center itself. These reactions will be discussed later on.
The timing of events heavily depends on the structures of the glycosyl donors, acceptors and
promoters. If the productive glycoside forming reactions proceed too slowly, numerous side
reactions imply the degradation of the labile glycosyl donor. However, under more vigorous
conditions, the acceptors can be also destroyed.
O
X
G
promoter
δ
O
β
A
G
OR
minor
β
O
O
G
A
OR
(*)
O
H
Glycosyl Donor
OR α
R = Non-participating group
(benzyl, azido, etc.)
A-OH = Glycosyl acceptor
(A = Aglycone)
major
α
O
G
RO O
A
O
G
orthoester
(reversible)
O
O
O
R
O
X
G
O
promoter
O
R
β
O
G
O
O
O
major
A
O
G
β
O
H
A
O
A
O
R
R
Glycosyl Donor
O
O
CO-R = participating group
(R = alkyl, aryl, etc.)
G
minor
α
A
O
O
R
O
H
G
OO
A
R
O
Scheme 2
(*) Participation of the solvent has a strong influence on the stereoselectivity (See, p. 15)
3. Choices, challenges and problems of the glycosidic bond
The success of a coupling reaction between two sugars depends on the reactivity of the donor
and acceptor, on the promoter, on the kind of substituents on both saccharide units and, of
1
Barresi, F.; Hindsgaul, O. “Glycosylation methods in oligosaccharide synthesis” Modern Synthetic Methods,
1995, 7, 281-330.
-6-
course, on the preferred selectivity of the reaction towards the α- or the β-anomeric form. The
experience of the person conducting the experiment also plays a role.
If we take the synthesis of a simple trisaccharide molecule as a target we can enumerate the
choices, challenges and potential problems listed in the following.
RO
O
O
X + HO
promoter
solvent
Y
Z
Z
O
RO
O
Z
O
O
or RO
Y
X = leaving group
R = protecting group
Y = potential leaving group
Z = participating or
non-participating group
RO
RO
O
Z
O
+ HO
O
O
α-linkage
Z
O
α,α-linkage
Z
O
Y
Z
Y
O
Z
O
Manipulate
if needed
Z
O
Z
β-linkage
promoter
solvent
O
O
Y
Z
or
Z
X
RO
O
Z
O
O
Z
O
O
Y
Z
α,β-linkage
Scheme 3
Choices
1.- Choice of X and Z in the donor
2.- Choice of Y and Z in the acceptor
3.- Choice of the promoter or catalyst
4.- Choice of solvent and temperature
5.- Choice of protecting groups
Challenges and problems
1.- Anomeric selectivity for 1,2-cis or 1,2-trans linkages.
2.- Site selectivity and reactivity of acceptor OH groups (e.g. axial, equatorial, primary; Dgluco, D-galacto, C-3, C-4, or others).
3.- Configuration, substituent, steric and electronic effect in the donor and acceptor (e. g. Dglucopyranosyl and D-galactopyranosyl donors with identical substituents sometimes give
different α/β ratios with the same alcohol acceptor).
4.- Stoichiometry relative to the ratio donor:acceptor equivalents.
5.- Selective activation of anomeric groups (if X, Y are orthogonal groups that is have
different reactivities), Y can be activated in the presence of X.
6.- Iterative glycosylation in a stepwise manner or by block synthesis
7.- Minimum manipulation of protecting groups
8.- Prospects for solid-phase oligosaccharide and automated synthesis
-7-
4.- Structures and reactivity of glycosyl donors and of glycosyl acceptors used in
oligosaccharide synthesis.
Structures of glycosyl donors
There are numerous glycosylation methods involving different glycosyl donors. The name of
the glycosylation method generally reflects the functionality of the glycosyl donor except for the
Fischer glycosylation that uses reducing sugars and the Köening-Knorr procedures that use
glycosyl halides as donors.
O
NH
O
O
L
O
SeAr
SR
CCl3
O
O
S
SEt
S
Glycosyl xantate
O
Thioglycosides
Trichloroacetimidates
Glycosyl halides
(L = F, Cl, Br)
O
S Ar
O
O
O
O
O
O
R
O
O
O O Orthoester
O
R
P
X
Glycosyl phosphorous
(R = Alkyl, O-alkyl,
X = O, S, lone pair)
Pentenoyl Glycosides
Anomeric acetate
O
3
O
1,2-epoxide
R
O
Pentenyl Glycosides
O
Glycals
O
3
O
O
N
N
Glycosyl sulphoxide Anomeric diaziridines
O
Selenoglycosides
OH
Reducing sugars
(R = OR', SR', CN)
vinyl glycosides
(R = H, Me)
R
Fig. 1 Structure of glycosyl donors used in oligosaccharide synthesis.
As a rule it is difficult to predict which glycosylation method will be the most suitable to
solve a certain problem. Nevertheless, there are some factors influencing the reactivity of
glycosyl donors that should be taken into account and that can be further used in the
optimization of an oligosaccharide synthesis.
Reactivity of Glycosyl Donors
The reactivity at the anomeric center depends to a large degree on the choice of the
protecting groups specially those on C-2. Glycosyl donors are then classified in two main
groups: armed donors (with an ether group on C-2) more reactive than disarmed donors (with
esters, amides on C-2).
-8-
Ester groups induce some positive charge at the anomeric
O
δ
X-G
Slow
center making the formation of the oxonium ion a slower
OBz
OBz
O
O
X-G
OBn
Fast
Fig. 2
O
process.
When identical protecting groups patterns are desired,
OBn
reactivity may be controlled by different leaving groups.
Both the nature of the heteroatom X and substituent G of the leaving group will affect the
reactivity. The configuration of the glycoside also influences its reactivity. Another element of
control occurs via the use of different promoters P for leaving groups activation. Finally,
sterical/torsional factors also have an influence. Fused rings resist flattening of the pyranose ring
during oxonium ion formation). As examples, butanodione and ciclohexanedioneacetals (BDA
and CDA methodologies) on C-3 and C-4, also reduce reactivity.
A modern glycosyl donor must has the following characteristics:
Accessibility, high stability toward protecting group manipulations and mild activation
conditions.
Reactivity of Glycosyl Acceptors
With regard to the reactivity of the acceptor, this depends on the nucleophilicity of the
hydroxyl groups in partially protected carbohydrates that in turn depends on their nature (1º
more reactive than 2º), their spatial orientation (equatorial more reactive than axial), the
conformation of the sugar ring (4C1 or 1C4) and the presence of other protecting groups in the
molecule.2 It can be generalised that electron-withdrawing groups diminish the reactivity of the
acceptor. In addition, the steric hindrance of the groups has an influence i.e. bulky groups at C-6
such as OTBDPS or OTBDMS or OPiv reduce the yield of a 1→4 glycosylation to a large
extent.
5.- Promoters, Solvents and Experimental Conditions.
The nature of the promoter, generally a Lewis acid, has an influence in the sense that it
favours the departure of the leaving group. In addition, its nature classifies the reactions as
homogenous and heterogeneous and this has implications for the stereochemistry.
The solvent also has an influence on the overall rate of the process and on the stereochemistry,
especially in the case of non-participating glycosyl donors. Anhydrous solvents are required to
avoid competition from water. Solvents of low polarity, such as dichloromethane or ether are
frequently used. Sometimes polar aprotic solvent such as acetonitrile or nitromethane are used.
2
a) “Relative reactivities of hydroxy groups in carbohydrates”, Haines, A. H. Adv. Carbohydr. Chem Biochem.
1976, 33, 11-109. b) “Modulation of the relative reactivities of carbohydrate secondary hydroxyl groups.
Modification of the hydrogen bond network”. Moitessier, N.; Chapleur, Y. Tetrahedron Lett. 2003, 44, 1731-1735.
-9-
On the other hand, some solvents may also form complexes with the intermediate sugar
oxonium cations affecting the orientation of the incoming O-nucleophile. For example, diethyl
ether enhances the formation of α-glycosides while acetonitrile favours the accumulation of βanomers. This is explained by the formation of an exocyclic complex with the solvents that
hinder the β and α faces, respectively.
The influence of the combination promoter/solvents on the stereochemistry will be
commented later on.
O
-E
Et-O
BnO
Me
-C
G
G
t
O
Et
O
Et
BnO
α-glycosidation
N
O
β-glycosidation
G
BnO
N
Me
Scheme 4
Experimental Conditions
The experimental conditions are very critical for the success of the reaction. Generally, the
use of extremely dry solvents, inert atmosphere and molecular sieves that can act as acid
scavenger are needed. Sometimes a non-nucleophilic base is also needed.
The order in which the reagents are added is also important in some cases.
The normal procedure of adding reagents (NP) is appropriate for less reactive disarmed
donors. The promoter (P) is added over a mixture of acceptor (A) and donor (D). For highly
reactive armed donors, the inverse procedure (IP) in which the donor is added over a mixture of
acceptor and promoter is the most convenient.
This can be rationalized as follows:
D+P+A
IP
NP
D.P
Decomp.
P
P
A
D.A
P.A
A
PA
D
Fig. 3
For a donor and acceptor with similar reactivities the NP procedure is commonly used. For a
termolecular reaction D + P +A, due to the nature of the reagents the reaction is expected to
occur through an association D.P and then interaction with A to obtain disaccharide D.A. For
highly reactive donors this strategy is less successful because the donor can decompose in the
- 10 -
presence of P before interacting with A. The IP procedure in which the complex A.P is first
formed and then reacts with the donor, solves the problem.
Example:
Schmidt, R. R.; Toepfer, A. Tetrahedron Lett. 1991, 32, 3353.
CCl3
O
Me
O
NH
AcO
AcO AcO
OBn
O HO
O
OBn
O
BnOOBn
NP: 43%
IP, 78%
OTBS
N3
AcO
Et2O, TMSOTf
BnO
OBn
Me
AcO
AcO AcO
O
O
AcO
OBn
OBn
O
O
OTBS
O
N3
Scheme 5
6. Anomeric control in chemical glycosylation. Methods for stereoselective formation of
glycosidic linkages
Types of anomeric linkages
The stereoselective introduction of the glycosidic linkage is one of the most challenging
aspects in chemical oligosaccharide synthesis. The anomeric linkages can be classified
according to the relative and absolute configuration at C-1 and C-2.
O
O
Z
OR
1,2-cis
2-D-glycero
OR
Z
Z
O
Z
1,2-trans
2-D-glycero
H
O
OR
OR
1,2-trans
2-L-glycero
1,2-cis
2-L-glycero
HO O
O
H
OR
2-deoxyglycosides
H
OR
OH
2-keto-3-deoxyulosonic acids
Fig. 4. Different types of glycosidic linkages
The 1,2-cis- and 1,2-trans-2-D-glycero series (allo-, gluco-, gulo- and galactopyranosides)
and the 1,2-cis and 1,2-trans-2-L-glycero series (altro-, manno-, ido- and talopyranosides). In
addition, some miscellaneous glycosidic linkages can be identified, including 2-deoxyglycosides
and 3-deoxy-2-keto-ulo(pyranosylic) acids.
6.1. Preparation of 1,2-trans-glycosides by neighbouring group participation
The nature of the protecting group at C-2 of the glycosyl donor is a major determinant of the
anomeric selectivity. A protecting group at C-2 that can perform neighbouring group
- 11 -
participation (disarmed donors) during glycosylation will give 1,2-trans glycosidic linkages.
Nucleophilic attack of the alcohol at the anomeric center of the more stable oxonium cation 3
originated by participation of the neighbouring after departure of the leaving group X, results in
the formation of a 1,2-trans-glycoside 4. Glucosyl type donors will give β-linked products while
mannosides will give α-glycosides.
O
O
1
G
O O
O
R'
O
G
G
O
R'
O
OH
X
G
O
R'
R' 3
2
OR
O
4
O
Scheme 6. Preparation of 1,2-trans-glycosides by neighbouring group participation
6.2. In situ anomerization for the synthesis of α-glycosides (Lemieux)
Lemieux and co-workers introduced this procedure in 1975 as a way of controlling the
anomeric selectivity in armed donors with non-assisting functionality at C-2. The reaction
conditions (e.g. solvent, temperature, and promoter) will determine the anomeric selectivity. The
in situ anomerization procedure results mainly in the formation of α-glycosides.
O
O
ROH
G
BnO Br
G
major
BnO OR
Et4N Br
Scheme 7
Lemieux discovered that the α-haloglucopyranoside is in equilibrium with the more reactive
β-halide and that the equilibrium is catalysed by halide ions derived from tetraalkylammonium
halides, and the reaction proceeds with inversion of a highly reactive β-halide with the alcohol
component via nucleophilic substitution.
O
G
Br
O
G
Br
BnO
2
BnO Br
1
Scheme 8
This reaction is thought to proceed through several intermediates (Scheme 9).
At equilibrium the proportion of the α-halide is relatively high. The β-halide is less stable
because of the de-stabilization as a results of the anomeric effect but reacts more rapidly than
the α-halide with an O-nucleophile.
- 12 -
O
ROH
G
O
Br
G
BnO Br
5
BnO
6
Br
O
BnO
Br
Slow
O
G
Et4NBr
G
O
O
OR
ROH
O
G
ROH
OR
O
G
BnO
O
β-glycoside
G
BnO
β-bromide
ROH
Fast
G
BnO
Br
α-bromide
BnO
BnO
O
G
G
BnO OR
α-glycoside
BnO OR
7
8
Scheme 9. Preparation of α-glycosides by in situ anomerization
To allow substitution of the β-halide, the C-1-halide bond, in order to be broken, must be
antiperiplanar to the electron lone pair of the ring oxygen.3 To establish such an arrangement, a
conformational change to the highly reactive boat-like intermediate is required. This makes
reaction of the β-halide fast. In the case of the α-halide a conformational change is not required
since the C-1 halide bond is already anti-periplanar to the ring oxygen lone pair and the
substitution of the α-halide is slow. It is clear that the equilibrium rate must be fast enough to
ensure that sufficient β-halide is continuously present. If the difference in reaction rate between
the α- and β-halides with the alcohol is large enough, α-linked O-glycosides are obtained as
major compounds or exclusively.
The reaction requires very reactive glycosyl halides (armed) and long reaction times, in
particular when the originally tetra-alkyl ammonium bromides are used as catalysts.
The in situ anomerization procedure has proven to be very useful. The use of other liofilic
promoters such as mercuric bromide, silver perchlorate and silver triflate make it possible to
carry out the reaction with even less reactive halides. However, the stereoselective outcome of
the glycosylations is very dependent not only on the reactivity of the catalyst, but also on the
reactivity of both the halide and the acceptor. Careful adjustment of the reactivity of the two
different components is essential in order to obtain satisfactory results.
3
Kirby, A. J. The Anomeric Effect and Related Stereoelectronic Effects at Oxygen. Springer Verlag, Berlin, 1983
- 13 -
6.3. Heterogeneous catalysis (Paulsen).
Glycosylation of α-halides in the presence of an insoluble silver salt proceeds mainly with
inversion of configuration and formation of the β-glycoside. In this case, the equilibration
between glycosyl halides is restricted because there is no nucleophile in the reaction mixture and
the reaction will therefore proceed with inversion of configuration. Silver silicate and silversilicate-aluminate have often been used as the heterogeneous catalyst. These catalysts have
proved to be valuable in the preparation of β-linked mannosides which can not be prepared by
neighbouring group participation or in situ anomerization.
BnO
BnO
O
O
G
BnO
O
ROH
G
G
9
OR
Br
Br
10
11
Ag
shielding α-face
Scheme 10. Glycosylation by inversion of configuration
However, the method only works well with very reactive halides and sufficient reactive
alcohol components. With less reactive components, significant proportions of the α-isomers are
obtained. β-Glycosides from glucose, galactose or fucose can also be prepared by the Paulsen
method, but it is usually more convenient to come along with strategies involving neighbouring
group participation.
6.4. Stereoselective preparation of α- and β-glycosides by participation of the solvent
The choice of the combination promoter/solvent plays a crucial role for the anomeric
stereocontrol of a glycosylation, especially when a non-participating group is at C-2 position.
In general, if any participating group is present at C-2, the glycosylation reaction follows a
SN2 pathway in non-polar solvents. The influence of the solvent under SN1-type conditions has
been extensively studied for ethers and nitriles.
O
Et-O-Et
BnO
BnOOR
promoter
O
+ ROH
L
α-glycosidation
G
(major compounds)
Me-CN
O
promoter
G
OR
β-glycosidation
BnO
Scheme 11
Ethers such as diethyl ether or THF favour the α-linkage while with acetonitrile, β-glycosides
are commonly obtained.
- 14 -
In diethyl ether, using strong acid promoters, the SN1-type reaction is favoured. Ethers
participate forming equatorial oxonium cations due to the reverse anomeric effect,4 which
favours thermodynamically α-glycosides.
O
O
promoter
G
BnO L
O
promoter
G
G
BnO
SN1
L
BnO
Et-O-Et
Et
O
O
G
BnO
reverse anomeric effect
ROH
Et
O
Scheme 12
G
BnO OR
α-glycosidation
The influence of nitriles, “The nitrile effect”, is more complex.5 Acetonitrile as polar solvent
favours an SN1 mechanism that implies the formation of an oxonium cation that is solvated with
preference at the α-face forming the kinetically controlled α-nitrilium-nitrile complex. This
complex finally renders the β-anomer by nucleophilic substitution by an alcohol.
On the other hand, the complex β-nitrilium-nitrile is thermodynamically more stable due to
the reverse anomeric effect, favouring the α-anomer. In any case, the complexation with the
nitrile increases the reactivity of the donor.
O
O
promoter
G
BnO L
O
promoter
L
G
G
BnO
BnO
Me-CN
SN1
β-glycosidation
S
O
O
N
G
Me
O
G
BnO OR
thermodynamic
control
4
5
N
α-glycosidation
ROH
S
BnO
S
BnO
ROH
G
Me
Scheme 13
S
O
G
OR
BnO
kinetic control
Lemieux, R. U. Pure. Appl. Chem., 1971, 25, 527.
Vankar, D.; Vankar, P. S.; Behrendt, M.; Schmidt, R. R. Tetrahedron 1992, 47, 9985
- 15 -
“The Nitrile Effect”
Scheme 14
For quite some time, there has been controversy with respect to the absolute configuration of
the intermediate α-glycosyl nitrilium ion. Trapping the intermediate nitrilium ion by 2chlorobenzoic acid gave the corresponding amide with α-configuration, thus confirming αnitrilium ions.6
O
G
Me-CN
O
R´-OH
G
BnO
O
G
BnO
N
O-R´
BnO
Me
Cl
COOH
O
O
G
G
BnO
BnO
Me
N
Ac
O
N
O
O
Cl
Cl
Scheme 15
Unfortunately this method gives low β-selectivity for mannosidases.
6
Ratcliffe, A. J.; Fraser-Raid, B. S. J. Chem. Soc., Perkin Trans I, 1990, 747.
- 16 -
6.5. Intramolecular aglycon delivery approach
This method has been applied with success to the synthesis of β-mannosides. In this method
the sugar alcohol (R´-OH) is first non-permanently linked to the C-2 position of a suitable
protected mannosyl donor via an acetal or silicon tether (Y = CH2 or SiMe2). Activation of the
mannose donor results in an intramolecular delivery of the alcohol in a concerted reaction
resulting in the formation of exclusively β-mannopyranosyl linkages.
HO
O
L R´-OH
X-Y-X
G
Y
OR´
O
O
L
G
Y
OR´
O
O
Y
O R´
O
O
G
HO
O
G
G
OR´
Scheme 16
Examples:
Stork , G. and La Clair, J. J. J. Am. Chem. Soc. 1996, 118, 247.
HO
O
BnO
BnO
BnO
O
+
S
HO BnO
O BnO
O
O
O
OC8H17
OBn
O
DMAP
78%
OC8H17
NPhth
Tf2O
S
Me2SiCl2, imidazole
NPhth
Ph
Si
BnO
BnO BnO
OBn
O
BnO
BnO
BnO
HO
O
O BnO
OBn
O
54%
Ph
OC8H17
NPhth
Tf-O-Tf
Scheme 17
Barresi, F. Hindsgaul, O. J. Am. Chem. Soc. 1991,113, 9367 and Synlett 1992, 759.
O
O
BnO
BnO
BnO
+
HO BnO
OBn
O
SEt
BnO
BnO BnO
I+
OBn
O
BnO
OC8H17
NPhth
NIS
SEt
TsOH
55%
NPhth
O
O
O
OC8H17
BnO
BnO BnO
HO
O
4-Me-DTBP
51%
O
OBn
O
BnO
OC8H17
NPhth
Scheme 18
- 17 -
8.- Common protecting groups used in oligosaccharide synthesis
Fig. 5
It is important to note, that in spite of the general approaches discussed above for
stereoselective control of the glycosidic linkage, other factors such as type of oligosaccharide,
leaving group at the anomeric center, protection and substitution pattern, promoter, solvent,
temperature, could have a major effect on the α/β selectivity.
It should be realized that there are no methods or strategies of general application for
oligosaccharide synthesis, which is one of its greatest difficulties. Nevertheless, convergent
multi-step synthetic sequences that give complex oligosaccharides consisting of up to 20
monosaccharide units are currently feasible by applying different strategies that will described
on Lesson 3.
- 18 -
Lesson 2. - Different procedures of glycosylation reactions by direct
activation
1.
2.
3.
4.
5.
6.
Köenings-Knorr method and related. Glycosyl Fluorides (Mukaiyama)
n-Pentenyl glycoside method (Fraser-Reid)
S-Glycoside methods (Lönn, Garegg, van Boom)
Phenylselenoglycosides
O-Alkylation and the trichloroacetimidate method (Schmidt)
Glycosylation with glycals (Lemieux, Thiem, Danishefsky)
Introduction
From a chemical point of view, the synthesis of oligosaccharides still presents an important
challenge to synthetic chemists in spite of major advances in the area. In this lesson we will
briefly review the main synthetic methods available for glycoside bond formation. Although
some methods for glycoside synthesis are more popular than others, there is no universal
protocol that can be applied to any combinations of donors and acceptors without consideration
of their substitution patterns, configurations, or position of the hydroxyl groups. All the choices,
challenges and potential problems that have been commented on in Lesson 1, are mostly
applicable to the various glycosylation methods.
Strategies for the assembly of sugars will be discussed in the next lesson.
1. Köenings-Knorr and related methods.
The Köenings-Knorr method uses glycosyl bromides and chlorides as donors in the
glycosylation reaction. It was first performed in 1901 and up until the mid-1980s, the method
and its numerous variants have been extensively used to prepare a wide variety of O-glycosides.
Insoluble promoters such as Ag2O and Ag2CO3 were initially used. Soluble catalysts
including HgBr2 and Hg(CN)2 (Helferich-Weiss, 1956) and AgOTf (Hanessian-Banoub, 1977),
were exploited as promoters. In the latter case, the reactions were sometimes performed in the
presence of tetramethylurea as acid scavenger.
Examples:
Hanessian, H.; Banoub, J. Methods in Carbohydr. Chem. Vol. 8, Whistler, R. L.; BeMiller, J.
N. Eds. Academic Press, New York, 1980, 247.
AcO
AcO
AcO
Ph
O
AcO
+
Br
O
O
HO
TfOAg, CH2Cl2
O
AcHN
OMe
Me2NCONMe2
82%
(based on
consummed ROH)
Scheme 1
- 19 -
Ph
AcO
AcO
AcO
O
AcO
O
O
O
O
AcHN
OMe
Betaneli, V.; Ovchinnikov, M. V.; Backinowsky, Kotchekov, N. K., Carbohydr. Res. 1980,
84, 211-214.
AcO
AcO
AcO
Ph
OAc
O
+
O
O
O
O
OAc
AcO
AcO
AcO
Hg(CN)2
O
O
HO Me
Br
O
O
O
Ph
O
O
O
O
MeCN
81%
O
(based on
consummed ROH)
O
O
OH
HO
HO
O
O
Me
O
O
HO
HO
O
O
HO
O
Me
HO
O
O
HO
OMe
OH
Scheme 2
In spite of the generality of the method there are several inconveniences that have limited its
use. The intrinsic instability of glycosyl halides, the requirement of at least an equimolar amount
(often up to 4 eq) of metal salts as promoters (frequently incorrectly termed as “catalyst”) and
problems concerning the disposal of waste material (e. g. mercury salts) have made the method
become less popular nowadays.
Other alternative methods of great interest have been developed.
1.1. Glycosyl fluorides (Mukaiyama)7
In 1981, Mukaiyama and co-workers introduced anomeric fluorides for the preparation of Oglycosides. The introduction of fluorine as leaving group is a good alternative to the KöeningsKnorr method due to the stability of the C-F bond. Glycosyl fluorides are easier to handle than
glycosyl chlorides or bromides. They are typically prepared from the anomeric acetates by
reaction with HF/py, from hemiacetals by reaction with DAST or from thioglycosides by
reaction with NBS/DAST.
Examples:
Hayashi, M.; Hashimoto, S.; Noyori, S. Chem. Lett. 1984, 1747.
BnO
BnO
BnO
HF- py
OAc -20ºC to 25ºC
OBn
80%
O
BnO
BnO
BnO
Scheme 3
7
For a review, Toshima, K. Carbohydr. Res. 2000, 327, 15-26.
- 20 -
O
BnO
α:β = 95:5
F
Posner, G. H. Haines, S. R. Tetrahedron Lett. 1985, 26, 5-9.
BnO
BnO
BnO
O
OH
OBn
DAST, THF
-30ºC to 25ºC
99%
BnO
BnO
BnO
O
F α:β = 1:7.7
BnO
Scheme 4
Nicolaou, K. C.; Dolle, R. E.; Papahatjis, D. P.; Randall, J. L. J. Am. Chem. Soc. 1984, 106,
4189.
AcO
AcO
AcO
O
SPh
NBS/DAST
CH2Cl2
-0ºC to 25ºC
70%
AcO
AcOAcO
O
100% α
F
Scheme 5
Because of the difference in halophilicity of this element compared with bromine and
chlorine, the glycosylation reactions require the use of other promoter systems besides silver
salts.
Mukaiyama and co-workers carried out the first reaction in 1981. In this case, 1,2-cis-αglycosides were predominantly obtained in high yields due to the anomeric effect.
Mukaiyama, T.; Murai, Y.; Shoda, S. Chem. Lett. 1981, 3, 431-432.
Scheme 6
Apart from SnCl2-AgClO4 (Mukaiyama, 1981), the following systems have been used:
TMSOTf (Hashimoto et al, 1984), BF3.Et2O (Kunz, 1985), Cp2ZrCl2-AgBF4 and Cp2HfCl2AgTfO/AgClO4 (Suzuki et al, 1989 and Mattheu et al, 1992), Cp2ZrCl2-AgClO4 (Matsumoto et
al, 1988), La(ClO4)3 (Kim et al, 1995 and LiClO4) (Böhm and Waldmann, 1995). The
promoters of wider application imply the use of lanthanide metals.
The glycosylations with anomeric fluorides follow the general principle as described for
bromides and chlorides. Apart from their enhanced stability, anomeric fluorides have not proven
to be superior to bromides or chlorides in terms of glycosylation efficacy.
- 21 -
Examples:
Mukaiyama, T.; Hashimoto, Y.; Shoda, S. Chem. Lett. 1983, 935-938.
Scheme 7
Takahashi, Y.; Ogawa, T. Carbohydr. Res. 1987, 164, 277-296.
Scheme 8
Wessel, H. P.; Ruiz, R. J. Carbohydr. Chem. 1991, 10, 901-910.
Scheme 9
Example:
In the total synthesis of NodRm-IV Factors:
Nicolaou, K. C.; Bockovich, N. J.; Carcanague, D. R.; Hummel, C. W. Even, L. F. J. Am.
Chem. Soc. 1992, 114, 8701-8702.
Nod Factors are the molecules signals involved in the symbiosis between legume plants and
bacteria of the genus Rhizobium. This symbiosis is responsible of the fixation of atmospheric
nitrogen in the roots of specific legume plants.
- 22 -
Structure and retrosynthetic analysis of Nod factors.
OTBDMS
OH
O
HO HO
a
OH
O HO
NH
d
b
c
OH
O
O HO
NHAc
O
OSO3
O HO
NHAc
O
-
PMBO
O
PMBO
F
OTBDMS
OMP
AcO
PMBO
NPhth
OH
O
F HO PMBO
NPhth
O
OMP
NPhth
NHAc
OH
O
O
Scheme 10
The key steps in the total synthesis imply glycosylation with glycosyl fluorides.
OMP
AcO
PMBO
+
F HOPBMO
NPhth
O
OMP
OBn
O
AgOTf, Cp2ZrCl2
OMP
CH2Cl2
NPhth
0º - 25º
O
AcO
PMBO
O
PBMO
NPhth
OBn
O
OMP
NPhth
NaOMe/MeOH
N
56 %
OMP
OMP
AcO
PMBO
O
HO
PMBO
O
O
PBMO
NPhth
OBn
O
OMP
NPhth
O
F
NPhth
OMP
AcO
PMBO
AgOTf, Cp2HfCl2
CH2Cl2
0º - 25º
O
PMBO
NPhth
N
60 %
OMP
O
O
PBMO
NPhth
OBn
O
OMP
NPhth
Scheme 11
Glycosyl fluorides are used together with thioglycosides in a double activation strategy. This
will be discussed in the next lesson.
2. n-Pentenyl glycoside method
This method, that uses pentenyl glycosides as glycosyl donors, was introduced by FraserReid in 1988. The activation of the leaving group is based on an electrophilic addition to the
double bond of the aglycone, followed by an intramolecular displacement by the ring oxygen
and eventual expulsion of the pentenyl chain to form an oxonium specie. Trapping with a
glycosyl acceptor, then leads to the desired glycoside.
- 23 -
G
E
E
O
O
G
R
E
O
O
G
R
E
O
O
G
R
R
Sugar-OH
O
G
O-Sugar
R
Scheme 12
The promoter of choice is any source of halonium ion. NBS or NIS alone or activated by
Lewis acid. NIS/Et3SiOTf is commonly used. Sometimes TfOH is also used. When using
halosuccinimides alone, the reaction is very slow, and often requires hours or days for
completion. A promoter of intermediate potency is IDCP (iodonium dicollidone perchlorate).
O
promoter
OH
+
G
X
O
G
NPG
Scheme 13
O
NPG
O
G
OCOPh
H
O
G
I+
OH
+
O
G
PhOCO Br
2,6-lutidine
Bu4NI
R
Sugar-OH
O
G
O-Sugar
R
I+
O
G
O
Ph
O
O
NPOE
Scheme 14
Preparation of n-pentenyl glycosides (NPGs) may be carried out following standard
procedures for preparing alkyl glycosides, including Fischer or Koenigs-Knorr glycosylations
with 4-pentenol.
When using perbenzoylated glycosyl bromides, reaction with 4-pentenol gives n-pentenyl
1,2-orthoesters (NPOEs) which can also serve as glycosyl donors. NPOEs are transformed into
- 24 -
NPGs through an acid-induced rearran-gement. The promoters of choice is NIS. Recently,8 an
efficient activation of NPOEs with NIS and lanthanide triflates (Yb(OTf)3) has been reported.
The advantage of using orthoesters is that they are stable to bases and so, several basepromoted protecting group transformations can be carried out before the acid-induced
rearrangement that converts NPOE to NPG.
Basically, both donors proceed mechanistically in the same way. They generate the same
intermediate that leads to the oligosaccharide.9
Scheme 15
NPOEs have the advantage over NPGs of the high stereocontrol observed due to the effective
shielding of the α (for D-Man) and β (for D-Glc) faces. Thus the reaction of benzoyl bromides
with 4-pentenol gave the NPOEs that show a high stereocontrol in glycosidic linkage formation
shielding the β and α faces of D-mannose and D-glucose that lead to α- and β-glycosides,
respectively.
8
9
Jayaprakash, K. N.; Radhakrishnan, K. V.; Fraser-Reid, B. Tetrahedron Lett, 2002, 43, 6953-6955.
Macha, M.; Schlueter, U.; Mathew, F.; Fraser-Reid, B.; Hazen, K. C. Tetrahedron 2002, 58, 7345-7354.
- 25 -
Example: Macha, M.; Schlueter, U.; Mathew, F.; Fraser-Reid, B.; Hazen, K. C. Tetrahedron
2002, 58, 7345-7354.
Conditions:
(i) PhCOCl, pyridine, DMAP; DCM; (ii) Ac2O, 30% HBr-AcOH(~85%); (iii) DCM, 2,6-lutidine, R′-OH or 4pentenol, Bu4NI; (iv) NaOMe, MeOH (89%); (v) NaH, BnBr, DMF (84%).
Scheme 16
Protecting groups influence the reactivity of pentenyl glycosides as donors. The so-called
armed-disarmed concept.
Example:
OBn
BnO
O
BnO
OPent
OBn
Armed
OH
+
O
OBn
OPent
AcO
AcO
OAc
Disarmed
IDCP
BnO
O
BnO
O
OBn
AcO
O
AcO
OPent
OAc
Scheme 17
Examples of glycosylations with NPOEs.
Recently, a strategy for Fully Inositol Acylated and Phosphorylated GPIs by the Synthesis of
a Malaria Candidate Glycosylphosphatidylinositol (GPI) Structure, has been reported using
NPOEs as donors.
Lu, J.; Jayaprakash, K. N.; Schlueter, U.; Fraser-Reid, B. J. Am. Chem. Soc. 2004, 126, 75407547.
They are anchored to the cell membranes and are connected to proteins via a
phosphoethanolamine linker. Hundreds of GPI-anchored proteins have been identified in
organisms ranging from archeabacteria to humans. They occur in all mammalian cell types.
They have diverse functions, including hydrolytic enzymes, adhesion proteins, complement
regulatory proteins, receptors, prion proteins, and antigens.
- 26 -
Retrosynthetic analysis:
O
P OBn
O
O
OH
O
HO HO
HO
Protein
NH2
V
(Manα1)
2Manα1
2Manα1
V
IV
O
O IV
HO HO
O
HO HO
4GlcNH2α1
6Manα1
III
6myoIno
II
I
III
OH
O
O
HO
O
Ph
O
I
H2N
HO
O
R
2
BnO
HO
O
II
HO
OCOR1
D-mannose
OH
OH
O
O
HO HO
O
2
1
O P OH
R3
OBnOBn
HO
O
O
O
myo-inositol
O
O
Scheme 18
O
Synthesis:
II
BnO
II
Ph
BnO
PMBOBnO
1
HOBnO
O
O
O
I
BnO
(i) protection
(ii) 2, NIS/Yb(OTf)3,
O
O
O
(i) change of Bz to Tf
(ii) N3TMS
OBn(iii) deprotection
O
(i)
I
N3 BnO
O
O
Ph
TBDMSO
O
O O
BnO BnO
, NIS/BF3.OEt2
O
(ii) manipulation of
OBn OBn
protecting groups
OBn
98%
III
HO
BnOBnO
BnOBnO
OBn
O
O
BnO
O
I
N3 BnO
O
O
IV
O
Ph
BnO
O
O O
BnO BnO
II
BnO
OBn
OR
O
BnOBnO
III
OBn
O
BnO
O
NaOMe/MeOH
R = Bz
R=H
AcOAcO
BnO
BnOBnO
OBz
V
O
HO HO
(i) deprotection
(ii)introduction of aminophosphate moiety
(iii)deprotection
(iv)introduction of the fatty acid
(v) reaction with glycerylphosphoamidite
O
O IV
O
BnOBnO
III
OBn
O
BnO
O
BnO
II
N3 BnO
O
O
HO
HO HO
(ii) manipulation of
protecting groups
O
OBn OBn
Protein
NH2
V
O
O IV
O
HO HO
(vi) reduction
O
P OBn
O
O
OH
O
III
OH
O
HO
O
HO
O
I
O
II
O
I
H2N
HO
O
R2
OBn OBn
O
R1, R2, R3 various fatty acyl groups
R3
- 27 -
O
O P OH
O
O
Scheme 19
AcO AcO
Ph
TrO
O
O O
, NIS/BF3.OEt2
I
N3 BnO
O
O
O
TrO
(i)
O
O
BnO
, NIS/BF3.OEt2
OBn OBn
II
O
OCOR1
OH
OH
3. - S-Glycoside methods
There are several methods in which the anomeric carbon is activated by groups having
sulphur in place of the exocyclic hemiacetal oxygen. The best known example of this type of
protection/activation group is the alkyl(aryl)thio group (thioglycosides). Oxidized forms of
thioglycosides, such as sulfoxides can act as glycosyl donors as well as other derivatives like Sxantates. We will focus our attention mainly on thioglycosides. Glycosyl sulfoxides will also be
considered.
3.1. Thioglycosides
The sulfur atom in a thioglycoside is a soft nucleophile and is able to react selectively with
soft electrophiles suchs as heavy metal cations, halogens, and alkylating or acylating reagents.
This fact make thioglycosides very versatile agents in carbohydrate chemistry. Additionally, the
hydroxy and ring oxygen atoms of carbohydrates are hard nucleophiles, which can be
functionalized with “hard” reagents, without affecting alkyl(aryl)thio function.
O
O
OH
HO
O
SR
HO
SR
R'O
O
R"OH
OR"
promoter R'O
Scheme 20
An electrophile activates the thioglycoside by producing intermediate sulfonium ions, which
then give rise to glycosylating carbocationic intermediates that react with the alcohol giving the
glycoside.
E
O
S
R'O
E
R
O
O
S
R'O
OBn
R
R'O
OBn
ROH
O
O
R'O
OBn
OBn
R
E
O
S
R'O
O
O
E
R
S
R'O
O
R
O
O
R
ROH
R'O
O
O
O
R
R
O
R'O
O
O
R
O
R
Scheme 21
Although this possibility was known for a considerable time (Bonner, 1948; Ferrier, 1973), it
has been since 1984 that it has been extensively explored.
In 1984 Lönn first reported the use of methyl triflate as the first efficient general promoter for
direct glycosylation with thioglycosides. MeOTf has disadvantages because it is toxic and in the
- 28 -
presence of slow reacting glycosyl donors, it can give rise to methyl ethers in addition to
glycosides. For this reason, other thiophilic promoters have been developed.
For example dimethyl(methylthio)sulfonium triflate, DMTST (Fugedi, Garegg, 1986),
NOFB4 (Pozsgay, Jennings, 1987/88), MeSOTf, MeSBr (Dasgupta, Garreg, 1988), PhSeOTf
(Ogawa, 1989), MeI (Reddy, 1989), NIS, TfOH (van Boon, Konradsson, 1990), IDCP
(Veeneman, van Boom, 1990), TBPA (Sinaÿ, 1990).
Me
N
I
Me
ClO4
Me
S
Me
S
Me
OTf
Br
3
DMTST
IDCP
Me
NH SbCl6
TBPA
N
I
Me
TfO
IDCT
Fig. 2
Iodonium dicollidine perchlorate (IDCP) is better replaced by iodonium dicollidine triflate
(IDCT), which has similar reactivity and which does not require the use of AgClO4 in its
synthesis. MeOTf, DMTST, NIS-TfOH and in particular PhSeOTf are all most efficient
promoters that produce fast reactions. Tris(4-bromophenyl)ammoniumyl hexachloroantimonate
(TBPA) differs from others in that its cation is radical, and as such produces radical cationic
sulfonium ions as glycosylating species from thioglycosides.
Regarding stereochemistry, the glycosylations with thioglycosides follow the general
principle as described for bromides and chlorides.
With regards to the preparation of thioglycosides, they can be grouped into three categories:
A. Acid-promoted Displacement at the anomeric center. This implies the synthesis from a
sugar derivative of a thiol in the presence of a Lewis acid.
Example: Ferrier, R.; Furneaux, R. Methods, Carbohydr. Chem. 1980, 8, 251.
AcO
AcO
OAc
O
OAc
OAc
PhSH
BF3/Et2O
71%
AcO
AcO
OAc
O
SPh
OAc
Scheme 22
B. Base-promoted Displacement at the Anomeric Center. This implies the synthesis by Snucleophilic displacement at the Anomeric Center
Example: Tropper, F.; Andersson, F.; Grandmaitre, C.; Roy, R. Synthesis, 1991, 734.
AcO
AcO
OAc
O
AcO
Br
PhS Na
Phase transfer
catalysis
AcO
AcO
81%
Scheme 23
- 29 -
OAc
O
OAc
SPh
C. Synthesis by preparation of a 1-thioglycoside followed by S-alkylation. Once prepared
the 1-thioglycoside, it is alkylated with an alkyl halide, often in situ. Although the total
number of steps is higher, the reagents are cheap and the yields are high throughout.
Example: Horton, D. Methods in Carbohydr. Chem. 1963, 2, 433.
S
OAc
O
AcO
AcO
H2N
AcO
AcO
OAc
O
AcO
AcO
acetone
AcO
Br
OAc
O
NH2
OAc
80%
SH
AcO
MeI
K2CO3 aq.
100%
NH2 Br
OAc
O
AcO
AcO
Diisopropyl
ethylamine
NH2
S
SMe
AcO
87%
Scheme 24
There are many examples of glycosylations with thioglycosides.
Example: The synthesis of part of the carbohydrate structural component of a glycoprotein
isolated from fucosidosis patients. Lönn, H. Carbohydr. Res. 1985, 139, 115-121.
OAc
O
AcO
AcO
O O
O
AcO OAc
Me
HO
OBn
SEt
NPhth
O
O
OBn
OBn
OBn
BnO OBn
BnO HO
O
OBn
O
OBn
O
BnO
O
OBn
OBn
61% MeOTf, Et2O
OAc
AcO
AcO
O
AcO
AcO
AcO
AcO
O
OAc
Me
BnO
OAc
Me
O
O
O
O
NPhth
O
OBn
O
BnO OBn OBn
BnO OBn
OAc
BnO
O
O
O
O
O
NPhth
O
O
OBn
OBn
OBn
OBn
OBn
O
OBn
O
OBn
β-D-Galp(1
4)
α-L-Fucp(1
3)
α-L-Fucp(1
3)
β-D-Galp(1
4)
β-D-GlcpNAc(1
β-D-GlcpNAc(1
Scheme 25
- 30 -
2)
2)
α-L-Manp(1
6)
α-L-Manp(1
3)
D-Man
Protecting groups influence the reactivity of thioglycosides:
Veeneman, G. H.; van Boom, J. H. Tetrahedron Lett. 1990, 31, 275
OBn
O
BnO BnO
SEt
OBn
armed
IDCP
+
HO BzO
OBz
O
BnO BnO
91%
disarmed
SEt
OBn
O
BnO
O
BzO
OBz
O
BnO BnO
armed
1. NaOMe
2. NaH/BnBr/Bu4NI
OBz
OBz
disarmed
SEt
OBn
O
BnO
O BnO
HO
OBz
O
BzO
disarmed
OBn
O
SEt
OBn
SEt
BnO BnO
OBn
O
BnO
OBz
IDCP
72%
O
OBn
O
BnO
BnO
O BzO
OBz
O
SEt
OBz
Scheme 26
3.2. Sulfinil glycosides: the sulfoxide method
The use of glycosyl sulfoxides as glycosyl donors, provides a new and powerful method for
chemical glycosylations, where a glycosyl sulfoxide (also called sulfinil glycosides) reacts with
a glycosyl acceptor in the presence of a promoter, to give a di- tri- or oligosaccharide.
O
O
S Ph
+
HO
O
OG
promoter
O
O
O
OG
promoters: Tf 2O, TMSOTf, TfOH
acid scavenger: DTBMP
Scheme 27
The promoter systems for these sulfinil glycosides are triflic anhydride (Tf2O) or
trimethylsilyl triflates in stoichiometric amount or triflic acid in catalytic amount. The reaction
is always carried out in the presence of an acid scavenger (diterc-butyl methyl pyridine).
Daniel Kahne first developed this method and was able to glycosylate very unreactive
hydroxyl groups as the C-7 hydroxyl group in a deoxycholic acid derivative.10 He used two
types of glycosyl donors with non-participant and participant protecting groups. Yields are good
with non-polar solvents. In the absence of a neighbouring group, the stereochemical outcome of
10
Kahne, D.; Walker, S.; Cheng, Y.; Van Engen, D. J. Am. Chem. Soc. 1989, 111, 6881-6882.
- 31 -
the reaction is strongly influenced by the solvent: The yield of the β-glycoside increases with the
polarity of the solvent (nitrile effect). With a C-2 participating group, the final product is all β.
Glycosyl acceptor
Glycosyl donor
Me
Me
Me
COOMe
OBn
O
BnO
BnO
OBn
OH
EtOCO
PivO
PivO
Conditions
O
S Ph
OPiv
O
O
S Ph
OPiv
Product ratio (yield)
toluene
α:β = 27:1 (86%)
CH2Cl2
α:β = 1:3 (80%)
acetonitrile
α:β = 1:8 (50%)
dichloromethane
all β (83%)
Scheme 28
The sulfoxide-glycosylation method is highly efficient with rather unreactive nucleophiles,
has potential for chemoselective glycosylations and is applicable to the synthesis of
oligosaccharides on solid supports. However, the highly reactive donors used in this method
make it impractical in some cases due to their decomposition.
One advantage of the sulfoxide method is its flexibility and wide scope. It has been
demostrated that using a standard set of conditions, it is possible to construct families of
oligosaccharides. As an example, the syntheses of the Lewis blood group of antigens: Lewis a,
Lewis b and Lewis x (Lea, Leb and Lex).
Example: Yan, L.; Kahne, D. J. Am. Chem. Soc. 1996, 118, 9239-9248.
The synthesis of Lea begins at -78° C with the coupling of sulfoxide 1 and acceptor 2, the
promoter is triflate anhydride and di-tercbutylmethylpyridine as acid scavenger.
Lea
β(1→3)
OPiv
PivO
O
O
PivO
S
Ph
+
Ph
O
O
HO
O
N3SPh
OPiv
1
2
OBn
OBn
OBn
Me
O
AcO OAc
PivO
O
O
O
AcO
O
N3
OAc
SPh
PivO
Tf2O
OPiv
Ph
O
O
O
N3
OPiv
O
DTBMP
CH2Cl2, -78°
83%
PivO
O
SPh
3
α(1→4)
Tf2O
Me
DTBMP
CH2Cl2, -78°
95%
AcO
OBn
OBn
O
O
OBn
O
HO
O
N3
OAc
AcO
S Ph
4
5
6
HO
OH
OH
Me
O
HO
HO OH
O
O
O
HO
OMe
O
AcHN
OH
Scheme 29
- 32 -
PivO
OAc
+
Lea
O
SPh
A β(1→3) glycosidic bond is formed. The same reaction conditions were used in the
coupling of acceptor 4, obtained after normal manipulation of protecting groups, and
fucosylsulfoxide 5. An α(1→4) glycosyl bond is now formed. Subsequent transformation gives
the final molecule.
Leb
BnO
OBn
β(1→3)
Ph
O
O
BnO
S
O
O
HO
+
O
Ph
OPiv
N3SPh
2
7
OBn
Ph
O
O
O
N3
OPiv
O
DTBMP
CH2Cl2, -78°C
77%
BnO
O
SPh
8
α(1→4)
α(1→2)
OBn
OBn
OBn
Me
O
PivO
BnO OBn
O
O
O
BnO
O
N
3
O
SPh
Me
O
OBn
BnO
BnO
Tf2O
BnO
OBn
OBn
Tf2O
Me
DTBMP
CH2Cl2, -78°C
O
O
82%
OBn
S Ph
+
PivO
OBn
O
BnO
HO
O
N3
OH
O
SPh
9
5
HO
OBn
10
OH
OH
Me
O
HO
HO OH
O
O
O
HO
OMe
O
N3
O
Me
O
OH
OH
HO
Leb
Scheme 30
For the synthesis of Leb, the same reaction gave the β(1→3) linked disaccharide 8, that was
transformed into acceptor 9 with two unprotected hydroxy groups. Double glycosylation with
fucosylsulfoxide 5 gives tetrasaccharide 10 with two new α(1→4) and α(1→2) linkages.
Subsequent transformations gave the final compound. The yields are always very good, from 77
to 95%.
Lex contains the same three sugars as Lea but they are linked in a different manner: the
position of galactose and fucose are reversed. The first coupling reaction, with the formation of
a β(1→4) linkage, proceed in a slightly lower yield, probably because the HO-4 is greatly
hindered by the pivaloyl and para-methoxybenzyl groups. The reaction of 5 and 13, under the
same conditions, gave the α(1→3) new bond.
- 33 -
Lex
β(1→4)
OPiv
PivO
PivO
HO
PMBO
O
O
S
PivO
Ph
+
O
OPiv
N3SPh
11
2
OPiv
PivO
Tf2O
PivO
O
DTBMP
CH2Cl2, -78°C
65%
AcO
O
AcO
Me
O
SPh
12
α(1→3)
OAc
AcO
O
O
PivO PMBO N
3
PivO
AcO
O
O
N3
OBn
OBn
OBn
Tf2O
O
Me
DTBMP
CH2Cl2, -78°C
83%
SPh
AcO
O
O
AcO
AcO
S Ph
O
OBn
BnO
OAc
+
OBn
AcO
O
O
HO
N3
5
SPh
13
14
HO
OH
O
HO
HO
O
Me
HO
O
O
O
OMe
NHAc
OH
Le x
OH
HO
Scheme 31
4. Phenylseleno glycosides
Anomeric phenylselenides are interesting glycosyl donors. The phenylseleno substituent
behaves largely like thioglycosides with respect to stability towards protecting group
manipulations and lability towards electrophilic reagents.
O
O
OH
HO
HO
O
SeR
R'O
O
SeR R"OH
OR"
promoter R'O
E
O
Se
R'O
E
R
O
O
Se
R'O
OBn
R
R'O
OBn
ROH
O
O
R'O
OBn
OBn
R
E
O
Se
R'O
O
O
E
R
Se
R'O
O
R
O
O
R
O
O
ROH
R'O
O
R
R
O
R'O
O
O
R
O
R
Scheme 32
Phenylseleno glycosides are more reactive than thioglycosides allowing chemoselective
glycosylations.
- 34 -
Example: Mehta, S.; Pinto, B. M. J. Org. Chem. 1993, 58, 3269-3276
BnO
OBn
O
BnO
O
SePh
BnO
BnO
OH
OBn
O
(79% α/β :3/1)
SEt
BnO
OBn
OBn
BnO
IDCP
O
+ BnO
O
BnO
BnO
SEt
OBn
BzO
EtS
OBz
OH
O
BzO
OBz
+ BzO
SePh
BzO
O
SEt
NIS
TfOH
79%
BzO
OBz
O
O
O
BzO
OBz
OBn
BzO
BnO
OBn
Scheme 33
Both C-2 acylated and benzylated glycosyl donors can be activated with AgTfO. The
glycosylation is quenched with the presence of tetramethylurea or collidine. Thioglycosides are
usually stable towards AgOTf, so orthogonal glycosylations are feasible.
Example: Mehta, S.; Pinto, M. Tetrahedron Lett. 1993, 32, 4435.
OH
SePh
+ BnO
BnO
O
Me AcO
SEt
OBn
AcO
OAc
K2CO3
85%
OH
AcO
AcO AcO
O
Phth
SePh
+ BnO
BnO
O
AgTfO
O
AgTfO
O
SEt
OBn
Me AcO
AcO
O
BnO BnO
OAc
AcO
AcO AcO
O
SEt
OBn
O
Phth
BnO BnO
K2CO3
O
O
SEt
OBn
Scheme 34
As AgTfO and bases such as tetramethylurea or collidine are frequently employed in
glycosylations with glycosyl halides, chemoselective glycosylations of glycosyl halides in the
presence of selenoglycosides are also possible.
Phenylseleno glycosides can be prepared from peracetylated sugars by reaction either with
phenylselenol, or from glycosyl halides by reaction with potassium phenyl selenoates or from
diglycopyranosyl diselenides by reaction with alkyl halides under reducing conditions.
Example: Mehta, S.; Pinto, B. M. J. Org. Chem. 1993, 58, 3269.
OAc
Me
AcO
AcO
O
OAc
SePh
PhSeOH
BF3.Et2O
84%, α:β = 3.7:1
Scheme 35
- 35 -
AcOMe
AcO
O
OAc
Example: Benhaddou, R.; Czernecki, S.; Randriamandimby, D. Synlett, 1992, 967.
BnO
BnO BnO
O
Br
Se
OBn
2
BnO
BnO BnO
NaBH 3CN
O
Se
OBn
Scheme 36
4. O-Alkylation and the trichloroacetimidate method (Schmidt)
4.1. O-Alkylation method
The anomeric oxygen of a sugar can be activated for a glycosylation not only by acids
(Fischer glycosylation) but also by bases. Upon treatment a hemiacetalic sugar with a base, the
generated anomeric oxide can be alkylated leading directly and irreversibly to a glycoside. This
process is called anomeric O-alkylation.
Schmidt, R. Angew. Chem. Int. Ed. Engl. 1986, 25, 212.
X
RO
RO RO
O
OH
Y
Base
X
RO
RO RO
X
X
RO
RO RO
O
Y
O
O
Y
O
RO
RO RO
O
Y
H
R'X
R'X
X
X
RO
RO RO
O
RO
RO RO
O
Y
O
OR'
Y
OR'
(kinetic control)
(thermodynamic control)
Scheme 37
In this procedure, some inconveniences should be considered: The equilibrium between the
two anomeric forms and the open-chain form gives three sides of attack and also, a base
catalysed elimination in the open chain form could become an important side reaction.
Therefore, the yield, the regioselectivity and the stereoselectivity of the anomeric O-alkylation
was not expected to be outstanding.
However, Schmidt and co-workers have described several good examples of this method
including glycosylation of unprotected sugars.
- 36 -
Examples:
This method has been applied in the synthesis of lactosyl esphingolipid, by reaction of
hemiacetalic lactose with sphingosine triflate. The yield is moderate and the selectivity strongly
depends on the temperature.
W. Klotz, R. R. Schmidt, J. Carbohydr. Chem. 1994, 13, 1093.
OAc
OAc
AcO
AcO
O
O
AcO
O
OAc
OAc
+
N3
OH
NaH
1,2-diethoxyethane
r.t.
49%, β:α= 95:5
OAc
OAc
AcO
AcO
O
O
AcO
O
OAc
N3
O
OTBDMS
OAc
TfO
OTBDMS
Scheme 38
Chelation control can also become a dominant factor in the determination of the α/β
selectivity. Example: Synthesis of KDO-α-glycosides of lipid A derivatives.
Rembold, H.; Schmidt, R. R. Carbohydr. Res. 1993, 246, 137-159.
Scheme 39
The anomeric hydroxyl group of KDO has a low reactivity because of the effect of the
carboxyl group. Formation of an amide that releases electrons and the formation of bulky
benzylidene acetals that promotes a boat-like conformation on the sugar ring make the reaction
of the anomeric oxygen with triflate 2 possible. The coupling is performed twice to give the
- 37 -
trisaccharide backbone that was further transformed into the lipid A analogue. The boat-like
conformation is stabilised by a chelating effect with the cation Na+ and the solvent.
4.2. The trichloroacetimidate method
Electron deficient nitriles are known to undergo direct and reversible base-catalysed addition
of alcohols to the triple bond system, providing O-alkyl imidates. The free imidates can be
directly isolated as stable adducts.
N + ROH
R3C-C
NH
base
R3C
OR
Scheme 40
The reaction of hemiacetalic sugars in the presence of a base with trichloroacetonitrile gives
the anomeric trichloroacetimidates. In this way, the anomeric oxygen atom has been
transformed into a good leaving group.11
O
OH
RO
Cl3C-C N
Base
RO
O
O
CCl3
NH
Scheme 41
Taking into account the equilibrium between both anomers and the enhanced nucleophilicity
of equatorial oxygen atoms (owing to steric effects and to the stereoelectronic kinetic anomeric
affect), the equatorial (β)-trichloroacetimidate is generated with preference or even exclusively
in a very rapid and reversible reaction. However, this product anomerizes in a slow basecatalysed reaction through retro-anomerization of the 1-oxide anion. Through a new
trichloroacetonitrile addition, the thermodynamically more stable axial (α)-trichloroacetimidate
is formed (thermodynamic anomeric effect).
O
O
RO
RO
OH
OH
Base
Base
BH +
O
O
RO
RO
O
O
+ BH
Cl3C-C N
Cl3C-C N
O
O
RO
RO
O
CCl3
O
CCl3
N H
B
NH
(thermodynamic control)
(kinetic control)
Scheme 42
11
Schmidt, R. R.; Kinzy, W. Adv. Carbohydr. Chem. Biochem. 1994, 50, 21-123.
- 38 -
The equilibration between the two trichloroacetimidates can be speeded up by stronger bases.
O
RO
Cl3C-C N
OH NaH or DBU
O
RO
O
R = Bn
O
Cl3C-C N
OH
O
RO
K2CO3
O
CCl3
Cl3C-C N
OH NaH or DBU
OR'
or K2CO3
RO
NH
O
RO
CCl3
O
RO
OR'
O
CCl3
NH
R' = esters, amides, imides
NH
Scheme 43
Thus, with different bases both O-activated anomers can be obtained in pure form and high
yield. However, NaH is appropriate for axial trichloroacetimidates while weaker bases such as
K2CO3 is appropriate for equatorial trichloroacetimidates.
Fig. 3
Concerning the glycosylation step, reaction of donor and acceptor under very mild acid
conditions leads to the corresponding glycoside in an irreversible manner. Acids, such as
BF3.OEt2 or TMSOTf are used in catalytic amounts. The proton liberated on the glycoside bond
formation reacts with the forming leaving group. This leads to a stable, non-basic
trichloroacetamide that provides the driving force of the reaction.
Example: Synthesis of lactosamine. Schmidt, R. R. University of Konstanz, unpublished
results.
AcO OAc
O
+ HOAcO
AcO
AcO
O
CCl3
NH
O
HO OH
AcO OAc
OAc
OR
BF3.OEt2
O
AcO
AcO
AcHN
leaving group + H+
Cl3C-CONH2
Scheme 44
- 39 -
AcO
O
AcHN
O
OAc
deprotection
OR
O
HO
HO
O
AcHN
HO
O
OH
OH
The stereochemical requirements are the same as in other glycosylation methods.
Other mild activating species, such as, AgOTf, have also been used.
Example: Robina, I.; López-Barba, E; Fuentes, J. Tetrahedron 1996, 52, 10771-10784.
OAc
AcO
OBn
O
O
NPhth
AcO
CCl3
+
HO
O
BnO
OBn
OMP
BnO
O
OBn
NH
O
AcO
AcO
OBn
OAc
OAc
O
O
O
NPhth
AgOTf, Cl2CH2, 60%
Stereoselectivity β, 100%
Procedure (IP)
OAc
AcO
BnO
O
NPhth
O
AcO
OBn
AcO
AcO
NPhth
NH
Ag
CF3SO3
+
C O
CCl3
ROH
Glycoside
OBn
OBn
OOBn
O
CCl3
Ag
HN
S. P. Douglas, D. M. Whitfield and J. J. Keprinsky,
J. Carbohydr. Chem., 1993, 12, 131.
O
OMP
OBn
Scheme 45
For the synthesis of a tetrasaccharide derived from GlcNAc where the difference in reactivity
between donor and acceptor is high, AgOTf has proved to be convenient because it activates the
departure of the leaving group more slowly, thus minimizing decomposition of the donor.
Summary
Activation of the anomeric center with trichloroacetonitrile
•
•
•
•
Convenient Base Catalyzed Trichloroacetimidate Formation
Controlled acces to α- and β-compounds by choice of the Base
Thermal stability of α- and β-trichloroacetimidates up to room temperature
If required, silica gel chromatography can be performed
Glycosyl transfer
•
•
•
•
•
•
•
•
Catalysis by acids (mainly Lewis acids) under very mild conditions
Irreversible reaction
Other Glycosidic bonds are not affected
Usually High Chemical yield
Reactivity corresponds to the halogenose/silver triflate system
Stereocontrol of Glycoside Bond Formation is Mainly Good to Excellent:
Protecting groups with Neighbouring Group Participation: 1,2-trans-Glycopyranosides
ƒ β-Glycosides of: Glc, GlcN, Gal, GalN, Xyl, Mur, 2-deoxy-Glc
ƒ α-Glycosides of: Man, Rha
Protecting groups without Neighbouring Group Participation:
•
Catalyst BF3.OEt: Inversion of anomer configuration
ƒ β-Glycosides of: Glc, GlcN, Gal, GalN, Xyl, Mur, GlcUA
ƒ α-Glycosides of: Man, Rha
•
Catalyst TMSOTf : Thermodynamically more stable anomer
ƒ α-Glycosides of: Glc, GlcN, Gal, GalN, Man, Fuc, Mur
- 40 -
The outstanding significance of the trichloroacetimidate method lies in the ability of glycosyl
trichloroacetimidates to act as strong glycosyl donors under relatively mild acid catalysis. This
has been demonstrated by its use in many laboratories all around the world. The efficiency of
the method makes it appropriate for use in solid-phase, as will be commented on in the next
lesson.
This method has not only been used in oligosaccharide synthesis, but also in the chemistry of
natural products where sugars are glycosylated to different moieties.
Example: Synthesis of Macroviracin D.
Mlynarski, J.; Ruiz-Caro, J.; Fürstner, A. Chem. Eur. J. 2004, 10, 2214-2222.
This is a new type of glycolipid with a rather intriguing structure isolated from Mycelicum
Streptomyces sp., that exhibits strong antiviral activity towards several viruses including HIV,
herpes, simple and varicella zoster. The synthesis of this compound implies three main reactions
that are indicated by A, B, C in the scheme.
Glycosylation with trichloroacetimidates in the presence of TMSOTf in MeCN gives the βanomer in all the cases, due to the participation of the solvent.
HO
HO
OH
O
HO
O
OH
HO
HO
O
O
O
O
O
HO
OH
HO
HO
HO O
O
OH
O
O
O
HO
OH
HO
O
O
O
O
OH
OH
OH
HO
O
OH
O
HO
OH
OBn
HO
Zn
O
H
O
OBu
t
O
BnO
BnO
2
BnO
OBn
1
2
OH
Scheme 46
- 41 -
3
O
CCl3
NH
6.
Glycosylation with glycals (Lemieux, Thiem, Danishefsky)
Glycals in oligosaccharide synthesis were first used by Lemieux in 1960s, by Thiem in 1980s
and since then, by Danishefsky and co-workers.
Glycals can be used as glycosyl donors in two modalities.
in situ
activation
Glycosyl
acceptor
O
E
O
O
OR
transformation
into a glycosyl donor
O
E
X
Glycosyl
acceptor
E
Scheme 47
In the 1st motif, in situ activation makes the glycal act as glycosyl donor by forming a nonisolable intermediate. In the 2nd motif, the glycal is first converted into a glycosyl donor through
different types of reactions (epoxidation, azidonitration or sulfonamide glycosylation). That is,
the glycal is precursor of a defined glycosyl donor.
The pioneer experiments that used glycals as glycosyl donors, were done by Lemieux and
Thiem who used halonium-mediated coupling to suitable acceptors. This particular reaction has
the tendency to give a trans-diaxial addition and provides a crucial route to α-linked
disaccharides having an axial 2-iodo function at the non-reducing end.
PO
PO PO
OP
O
I
RO RO
PO PO
OP
O I
PO PO
I
O
Halo-glycosylation
O
OH
O
RO RO
OR
OR
O
OR
OR
Scheme 48
Because the displacement of an axial iodine atom has proven to be very difficult, azaglycosylation of glycals has been investigated with the idea of preparing glycosides of 2acylaminosugars.
Azidonitration with CAN/NaN3 was studied by Lemieux and constituted an important
advance at the time, nevertheless the conversion of the nitro-azido compounds into
oligosaccharides has not been fully optimized with regards to the yield and stereoselectivity.
- 42 -
glycosyl donors
OP
OP
CAN/NaN3
PO PO
O
ONO2
O
PO PO
NHAc
N3
I
PO
PhSO2NH2/IDCP
PO PO
O
PO PO
OP
O
Base
PO
PO PO
O
acceptor
PO PO
O
PO PO
PO
PO PO
Diisopropylidenegalactose
OP
acceptor
N
SO2Ph
NHSO2Ph
[O]
Azidonitro-glycosylation
OR
PO
O
O
PO PO
PhO2SHN PO
OH
PO
O
acceptor
PO PO
OH
O
O
O
HO
PO
PO PO
O
O
Sulfonamido-glycosylation
O
O
O
O
Aza-glycosylation
O O
O
HO PO
1,2-Anhydrosugar-glycosylation
O
PO
Scheme 49
Other procedures, such as iodo-sulfonamidation developed by Danishefsky, have been used
with more success for the synthesis of 2-acylamino oligosaccharides.
This method implies a trans-diaxial addition of an N-halobenzene sulfonamide to a glycal
followed by a base treatment that gives an intermediate that reacts with any acceptor, for
instance, another glycal, furnishing glycosides of benzenesulfonyl glucosamine derivatives:
sulfonamido-glycosylation.
While iodo-glycosylation and sulfonamido-glycosylation are rather good methods for the
conversion of glycals in various glycosides, the 1,2-anhydro sugar glycosylation provides a
general method for converting glycals into common oligosaccharides of glucose, mannose and
galactose in a high stereocontrolled manner. Once the glycal is converted into the 1,2-oxirane, it
may react with several acceptors leading to disaccharides. This method has been the most
widely used for the rapid assembly of oligosaccharides, and is appropriate for solid-phase
synthesis.
Protecting groups influence the reactivity of glycals as donors. The armed-disarmed concept
that prevails in pentenyl glycosides and thioglycosides is also applied here.
Example: Friesen, R. W.; Danishefsky, S. J. Tetrahedron 1990, 112, 8895
BnO BnO
OBn
O
+
HO BzO
OBz
O
I
BnO
I
58%
O
BnO BnO
O
BnO BnO
OBn
O
+ BzO HO
OBz
O
BnO
I
76%
- 43 -
BzO
I
O
BnO BnO
BzO
Scheme 50
OBz
O
OBz
O
O
When a benzylated glycal is made to react with benzoylated glycal no self-condensation is
observed and only one product is obtained derived from the more reactive glycal acting as
donor.
With regards to 1,2-anhydro sugars, the method was able to be applied when it was
discovered that glycals react smoothly with 2,2-dimethyldioxirane prepared as a solution in
dichloromethane, giving 1,2-anhydro sugars in good yields. The stereoselectivity of the
epoxidation highly depends on the type of protecting groups and on the steric hindrance of the
substituents.
Examples: Danishefsky, S. J. ; Halcomb, R. I. J. Am. Chem. Soc. 1989, 111, 6661.
DMDO
OBn
O
BnO BnO
OBn
O
O
O
CH2Cl2
BnO BnO
MeOH
BnO BnO
OBn
O
OMe
OH
O
α:β = 20:1
TBSO
OTBS
O
TBSO
TBSO
O
CH2Cl2
Ph
OTBS
O
O
only α
TBSO
O
O
O
O
TBSO
O
O
O
CH2Cl2
Ph
O
O
O
O
O
Ph
Ph
O O
TBSO
O
O
O
CH2Cl2
O
TBSO
O
α:β = 1:1
TBSO
β >>>α
yields, 90 to 100%
Scheme 51
The 3,4,6-tri-O-benzyl-D-glucal gives the epoxide in quantitative yield. Its solvolysis gave
the corresponding methyl glycoside with a stereoselectivity of 20:1 in favour of the α-isomer.
With resident acetyl protecting groups, the stereoselectivity of the epoxidation is much reduced.
TBS protecting groups or acetals also give high stereoselective epoxidations. Steric
hindrance also has an influence. Reaction of TBS-protected galactal gives stereoselectively the
α-epoxide, while the presence of an axial substituent at C-3 on the glycal promotes a quite
selective epoxidation from its β-face. On the other hand, the gulal configurated glycal with
hindering substituents on both faces of the double bond gave a 1:1 mixture of epoxides.
Examples:
Synthesis of Kijanimycin: Thiem. J.; Köpper, S. Tetrahedron 1990, 46, 113.
Halo-glycosylation has been mainly applied to the synthesis of 2-deoxy sugars due to the
inconveniences that the substitution of an iodine atom from the C-2 position generally offers.
NIS promoted glycosylation of glycals followed by reduction with H2/Pd and manipulation
of protecting groups furnished the desired oligosaccharide (Scheme 52).
- 44 -
OH OBn
BnO
BnO Me
O
BnO Me
NIS
+
OBz
O
MPMO Me
O
1. H2/Pd/C
2. NaOMe
OBn
O
MeCN, r.t. MPMO Me
OBz
O
O
MPMO Me
OBn
3. NIS, MeCN,
r. t. OBn
I
O
DDQ
Me
BnO
O
AcO Me
O
AcO Me
48% (α anomer)
O
O
I
BnO
O
DDQ
HO Me
OBn
O
O
O
OMPM
BnO
Me
BnO
O
AcO Me
OMPM
Cl
AcOMe O
O
Me
BnO
OBn
O
AcO Me
AgOTf
O
O
Me
O
O
O
AcO Me
I
I
HO
O
OH
O
HO Me
O
O
O
Me
OH
Kijanimycin
HO Me
O
Me
HO
O
Scheme 52
A similar method has been applied for the synthesis of Avermectine
Example: Danishefshy S. J.; Selnick, H. G. ; Armistead, D. M.; Wincott, F.E. J. Am. Chem.
Soc. 1987, 109, 8119.
OMe
OMe
O
HO Me
OMe
O
NIS
+
O
OMe
O
Me
AcO
O
OMe
O
Me
OMe
OMe
AcO
AcO Me
O
Me
O
Me
OMe
I
1. NIS, 64% (α anomer)
2. Bu4SnH-AIBN, 78%
3. LiEt3BH, 97%
66% (α anomer)
Me
Me
MeO
O Me
AcO
Me
O
Me
Me
H
O
O
OMe
Me
O
O
HO
Me
O
H
OMe
Avermectin 1α
Scheme 53
Example: Total synthesis of Tumor-Related Antigens N3, isolated from human milk. Its
composition depends on the blood type of the lactating mother.
Kim, H. M.; Kim, I. J.; Danishefshy S. J. J. Am. Chem. Soc. 2001, 123, 35-48
- 45 -
Retrosynthesis:
OH
HO
HO
OH
O
Me
HO
O
HO
OH
OH
O
O
O
O
OH
O
PO
O
HO
OH
OH
OH
OP
PO
O
PO
OP
O
Me
PO
OH
O O
OP
OP'
O
O
O
PO
OH
PO
O
O
O
OP
O
OP
PO
O
HO
O
PO
OP
Aza-glycosidation
O
F
PO
OP'
OP
PO
O
OP
OP
O
OP
OP
PO
OP
O
O O
OH
PO
HO
Me
NHP
O
OP
PO
Aza-glycosidation
HO
Me
O
O
OP
O
OP
O
O
OP
O
O
OP
PO
OP
O
NHP
O
OP
PO
PO
PO
OP
OP
PO
Difucosyllacto-N-hexaose
OH
HO
OH
HO
Me
O
PHN
O
O
PO
O
O
O
OP
OP PO
O
NHAc
HO
Me
OP
O
Me
OH
O O
HO
PO
O
AcHN
OH
OH HO
OP
O
O
O
O
OP
PO
OH
O
P = Generalized Protecting Group
P'= C-6 Protecting Group
O
P"O P"O
PO
O
Me
OP
OP
PO
P" = P or H
Scheme 54
Synthesis
O
+
O
DMDO
CH2Cl2
O
HO BnO
Lev
HO
OTIPS
TIPS
O
O
MPG : manipulating Protecting groups
MPG
OTIPS
O
O
HO
O
BnO
OTBS
7
5
4
NaMeO
TIPS
O
O
O
O
O
F
O OBn
OBn
BnO
AgClO4
O
HO Ph SiO
3
MPG
O
Me
1
Fucosylation
BnO
O
OBn
BnO
HO
NaMeO
MPG
MeOH
F
O OBn
OBn
BnO
O
Me
AgClO4
MPG
OBn
BnO
11
O
O
OBn
14
MeOTf
NHSO2Ph
OBn
OBn
MeOH
O
8
OTIPS
O
O
BnO
OH
MeOTf
TIPSO
TIPSO
PMBO
I
O
IDCP
O
Me
PhSO2NH2
MPG
OBn
12
BnO
aza-glycosylation
OBn
OBn
O
O
SEt
O
EtSH
LHMDS,
NHSO2Ph
O
DMF
Me
OBn
NHSO2Ph
OBn
14
BnO
PMBO
13
HO
9 +
I
9
OBn
O
HO O
O
NHSO2Ph
O
Me
OH
HO
TIPSO
O
OH O
O
O
BnO
O
BnO
6
TIPS
OTIPS
OTIPS Me
O
Me
aza-glycosylation
OBn
TIPSO
O
OH O
O
O
3
O
HO HO
OBn
O
IDCP
PhSO2NH2
OBn
O
7+8
O
Me
O
O
O
OH O
TIPS
O
OTIPS
O
O
DMDO
CH2Cl2
OTIPS
2
+
OTIPS Me
TIPS
O
TIPSO
O
HO
O
O
NH
PhSO
2
O
Me
OBn
MPG
OBn
BnO
OTIPS
O
HO
O
O TIPSO
O
O
O
O
O
OH O
NHSO2Ph OH
O OBn
Me
OBn
BnO
OH
O
O TIPS
O
O
SEt
O
OH
MeOTf
OTIPS
O
BnO
O
Scheme 55
- 46 -
HO
MPG
HO
OH
OH
O
Me
OH
HO
OH
O
HO
HO
O
Me
OH
HO
O O
O
O
O
O
AcHN
OH
OH
O
HO
OH
O
O
NHAc
OH
OHO
OH
Difucosyllacto-N-hexaose
O
OH
OH
Example: Synthesis of a branched oligosaccharide fragment of a complex Saponin:
Desgalactotigonin.
Randolph, J. T.; Danishefsky, S. J. J. Am. Chem. Soc. 193, 115, 8473-8474.
O
OH
HO
O O
HOHO
O
O
OH
O
O
HO
OH
Me
Me
OH
HO
HO HO
O
H
Fig. 4
OH
RO
1: desgalactotigonin (R=tetrasaccharide)
2: tigogenin (R=H)
H
The strategy consists on the preparation of a glycal epoxide that reacts as donor with a
glycosyl acceptor leading to a C(1)-O-sugar, with one hydroxyl group at C-2. This derivative
acts as glycosyl acceptor when it reacts with a glycosyl donor furnishing a branched
trisaccharide.
OP
OP
O
PO PO
O
PO PO
O
4
3
GA
OP
OP
O
PO PO
OSugar
OSugar
6
O
PO PO
GD
OSugar
Scheme 56
OH
5
GA : Glycosyl acceptor
GD: Glycosyl donor
This idea is exemplified in the following route:
BnO BnO
O
O
DMDO BnO BnO
CH2Cl2
Ph
O
HO O
O
O
Ph
O
BnO BnO
ZnCl2
O
O
O
O
MPG
BnO BnO
OH
THF
Ph
O
O
O
O
O
OBn
O
Zn(OTf)2
MPG : manipulating Protecting groups
O
OTIPS
O
O
O
O
O
DMDO
CH2Cl2
OTIPS
O
HO
tigogenin
O
OBn
MPG
O
BnO
tigogenin
OBn
O
OH
HO
O O
HOHO
BnO BnO
Ph
O
OBn
O
O
O
O
O
OH
BnO
OBn
O
OBn
O
F
OBn
Sn(OTf)2
BnO BnO
tigogenin
MPG
O
HO
OH
HO
HO HO
O
O
O
OH
O
O
H
OH
OH
H
OBn
Me
O
Scheme 57
- 47 -
Me
- 48 -
Lesson 3. Synthetic Strategies for the Assembly of Oligosaccharides
1.
2.
3.
4.
5.
6.
The pioneer linear glycosylation strategy
Convergent block synthesis
Selective and two-Stage Activation and Orthogonal Glycosylation strategy
Chemoselective Glycosylation Reactions
One-pot multistep glycosylations
Solid-phase oligosaccharide synthesis
Introduction
In this lesson, we are going to comment on different strategies for the assembly of
oligosaccharides with the idea of achieving the most efficient total synthesis of a complex
oligosaccharide. We will consider several approaches that allow the convenient assembly of
complex oligosaccharides from properly protected building block units involving a minimum
number of synthetic steps.
1.-The pioneer linear glycosylation strategy
In the pioneer linear glycosylation strategy, monomeric glycosyl donors have to be added to
a growing saccharide chain. Each step requires manipulation of protecting and leaving groups
which increases the number of reaction steps considerably. This fact, together with its low
convergence, makes this linear strategy the least efficient for the synthesis of complex
oligosaccharides. It has been used with glycosyl halides that require drastic reaction conditions
for their preparation and, in consequence, is incompatible with complex oligosaccharides.
2. - Convergent block synthesis
It is applicable for glycosylation methods in which the donors are formed under mild
conditions, are stable enough to be purified and stored for a considerable period of time, and are
able to undergo the glycosylation step also under mild conditions with high yield and high α/β
stereoselectivity. Trichloroacetimidates, thioglycosides, glycosyl fluorides and glycals have
been extensively used in block synthesis because they fulfil these requirements.
In a convergent glycosylation strategy most of the synthetic effort is directed towards the
preparation of monomeric glycosyl donors and acceptors. The assembly of these units to an
oligomer should involve the minimum number of synthetic steps and each synthetic step should
proceed with high stereoselectivity and high yield. Furthermore, an efficient synthetic
convergent strategy should make optimal use of common intermediates and oligosaccharide
building blocks.
- 49 -
Example: Several high-mannose and hybrid types of oligosaccharides have been recently
prepared as synthetic Carbohydrate-Based HIV Antigens using this strategy.
Dudkin, V. I.; Orlova, M.; Geng, X.; Mandal, M.; Olson, W. C.; Danishefsky, S. J. J. Am.
Chem. Soc. 2004, 126, 9560-9562
Gp120 carbohydrates can be used as antigens for eliciting broadly neutralizing immune
response. This idea has gained recognition after the structural determination of 2G12 antibody
epitope, isolated from long-term survivor of infection. This antibody is able to neutralize a wide
spectrum of HIV isolated in vitro and to protect macaques from SIV. The envelope glycoprotein
gp120 of HIV interacts sequentially with the cellular receptors CD4 and a member of the
chemokine co-receptor family.
HO
OH
O
HOHO
HO
High-mannose type glycopeptides
D-mannose pentasaccharide branch
O
O
HOHO
OH
O
O
HO O
OO
HO O
core D-mannose chitobiose trisasaccharide
O
OH
O
HO
HO
O H
N
O
HO
OH
NHAc
HO
NHAc
OH
O
OH
H2N-Cys- Asn-Ile-Ser-Arg-NH2
O
OH
1
O
O
SR
OH OH
O
OH
OH
HO
OH
D-mannose trisaccharide branch
OH
OH
OH
OH
HO
HO
HOHO
OH
O
HO OH
HO HO
HO
HO
O
HO
OH
HO
HO
O
HO
O
OHO
Hybrid type glycopeptides
O
HO O
HOO
D-mannose
trisasaccharide
branch
O
OHO
OH
O
O
HO O
O
core D-mannose chitobiose trisasaccharide
OH
O
HO
OHO
O
NHAc
6
O
HO
OHO
O
H
N
O
NHAc
H2N-Cys- Asn-Ile-Ser-Arg-NH2
SR
NHAc
O HO
D-lactose -D-mannose
trisasaccharide
branch
Fig. 1
The synthesis of high mannose oligosaccharides has been carried out by a convergent block
synthesis using thioglycosides 2 and 3 as donors that were coupled to the core D-mannose
chitobiose trisaccharide acceptor through the stereoselective formation of α(1→6)
and α(1→3) linkages, respectively giving the free glycan (Man9(GlcNAc)2). On its side, the
synthesis of the core trisaccharide has been carried out from glycal 7 by iodosulfonamidation
and reaction with 3,4-di-O-benzylglycal, to give the glycal disaccharide 6 that gave 4 by
- 50 -
iodosulfonamidation, manipulation of protecting groups and glycosylation with phenyl sulfinil
glycoside 10. [Dudkin, V.Y.; Miller, J. S.; Danishefsky, S. J. Tetrahedron Lett. 2003, 44,
1791]. Formation of the corresponding glycosyl amine in glycan Man9(GlcNAc)2 followed by
aspartylation with 5 gave the target glycopeptide 1.
BnO
High-mannose type glycopeptides
OAc
O
BnO BnO
BnO
BnO BnO
HO
HOHO
HO
OH
O
O
O
HOHO
O
HO O
HO O
(A)
OH
O
OO
O
HO O
HO
OH
HO
OH
O
(B)
OH
O
OH
O
O
OH OH
O
OH
OH
HO
1
OH
OH
OH OH
OH
D-mannose
trisaccharide branch
BnO
BnO O
O
O
O
OBn
O
BnO
SPh
2
OH
O
HO
OHO
BnO
OBn
OBn
OBn
OBn
Fmoc-HN-Cys-Asn-Ile-Ser-Arg-NH2
(C)
(D)
(D)
SSBut 5
O H
N
O
HO
O
NHAc
OHO
NHAc
H2N-Cys- Asn-Ile-Ser-Arg-NH2
O
BnOBnO
8
BnO
core D-mannose
chitobiose trisasaccharide
O
7
O
D-mannose
pentasaccharide branch
BnO
AcOBnO
O
O
OBn
O
Ph
O
PMBO O
SOPh
9
(C)
O
(B)
SR
OAc
O
BnO
BnO
BnO
BnO
BnO
Ph
O
HO O
O
O
BnO BnO
BnO
BnO BnO
OBn BnO
O O
BnO
BnO BnO
BnO
O
O
BnO
PhSO2NH
PhSO2NH
OTBS
4
O
O
O
O
10
O
OO
BnO
HO BnO
SEt
3
O
BnO
O
PhSO2NH
O
BnO
BnO
BnO
BnO BnO
O O
BnO
O
O
O
OBn
6
11
BnO
AcOBnO
O
7
Synthesis
7
iodosulfonamidation +
3,6-di-O-benzylglycal
(i) iodosulfonamidation
6 (ii) Manipulation of P.G.
(iii)Glycosylation with 9
4
(i) glycosylation with 3
(ii) deprotection
(iii) glycosylation with 2
(iv) global deprotection
Man9(GlcNAc)2 (i) amination
(free glycan)
(ii) aspartylation
with 5
Scheme 1
3.- Selective and Two-Stage Activation and Orthogonal Glycosylation strategies
Notwithstanding the attractive features of the above mentioned block synthesis, the
conversion of a common building block into a glycosyl donor requires several manipulations at
the anomeric center presenting the drawback of the removal of the anomeric protecting group
followed by the introduction of a leaving group, which can be a serious problem when
performed on larger fragments. The selective and two-stage activation strategy solves this
problem. In it, two types of anomeric leaving groups one obtained from the other, and one type
of activation is used.
In 1984, Nicolaou and co-workers described the glycosylation strategy that is outlined in
Scheme 2. Glycosylfluorides and thioglycosides are used. This two-stage strategy is convergent
- 51 -
1
and minimizes the number of manipulations, which have to be executed at the oligosaccharide
stage. Attractive features of the strategy are:
(i)
The stability of thioglycosides under many different chemical conditions.
(ii) The ease of activation of thioglycosides by conversion into glycosyl fluorides.
(iii) The high efficiency of glycosyl fluorides in glycosidic bond formation.
(iv) The excellent behaviour of thioglycosides as glycosyl acceptors.
O
RO
O
DAST
NBS RO
SPh
OR1
O
F
HO
OR1
activation
stage 1
AgClO4
SnCl2
activation
stage 2
O
O
RO
DAST
NBS
O
O
SPh
OR2
Deprotection
oligosaccharide
O
O
HO
OR1
O
O
F
OR2
OR1
SPh
OR2
HO
Glycosyl donor
O
SPh
OR2
OR1
Glycosyl acceptor
coupling
Higher
oligosaccharide
Scheme 2
Example: Synthesis of Rhynchosporides III
Nicolaou, K. C.; Dolle, R. E.; Papahatjis, D. P. J. Am. Chem. Soc., 1984, 106, 4189-92.
OTPS
O
AcO BnO
OTPS
BnO
F
OH
AcO BnO
AcO
AgClO4-SnCl2
CH2Cl2. -15ºC
O
BnO
BnO
AcO
O
BnO
SPh
O
O
BnO
BnO
SPh
DAST-NBS
CH2Cl2. 0º
-15ºC
TBAF-THF
0º
-15ºC
OH
OTPS
AcO
O
O
AcO BnO
BnO
BnO
BnO
O
AcO BnO
O
AcO
O
O
BnO
BnO
SPh
BnO
F
A
AgClO4-SnCl2
CH2Cl2. -15ºC
85%
OTPS
AcO
AcO
O
BnO
BnO
OTPS
O
AcO BnO
O
BnO
O
AcO BnO
O
BnO
BnO
1.- DAST-NBS
CH2Cl2. 0º
-15ºC, 85%
O
BnO
O
AcO BnO
A AgClO4-SnCl2
2.- CH2Cl2. -15ºC, 66%
O
BnO
AcO
O
O
AcO BnO
BnO
AcO
O
BnO
O
BnO O
O
O
HO BnO
BnO
BnO
SPh
Scheme 3
- 52 -
O
BnO
O
AcO BnO
O
BnO
SPh
Example: Synthesis of LeX fluoride:
Nicolaou, K. C.; Dolle, R. E.; Papahatjis, D. P. J. Am. Chem. Soc. 1990, 112, 3693
CAO OPiv
O
CAO
AcO
F
CAO
AgClO4-SnCl2
CH2Cl2. -15ºC
OBn
O
CAO
72%
O
HO AllylO
OPiv
1. H2Ru(PPh3)4, EtOH
then TsOH, MeO, 25ºC, 86%
OBn
O
O AllylO
AcO
F
SPh
2. AgClO4-SnCl2, Me
Et2O. -30ºC, 87%
PhthN
SPh
O
OBn
OBn
BnO
PhthN
CAO
OPiv
OBn
O
O
O
CAO
O
SPh
-15ºC
1. DAST-NBS, CH2Cl2, 0º
2. H2, Pd(OH)2/C, EtOH-EtOAc, 25ºC
3. Ac2O, DMAP, 2,6-lutidine, 25 ºC, 84%, 2 steps
CAO
Me
O
OAc
O
O
O
CAO
PhthN
AcO
OPiv
PhthN
O
Me
OBn
OBn
BnO
F
O
AcO
OAc
OAc
AcO
Scheme 4
Another two-stage activation strategy reported employs anomeric sulphoxides as donors and
thioglycosides as acceptors. The latter can be converted into sulfinil glycosides by oxidation.
Example: Khiar N.; Martin-Lomas, M. J. Org. Chem. 1995, 60, 7017.
OH
BzO
OBz
O
BzO
BzO
SPh
MCPBA
OBz
OBz
BzO
BzO
BzO
O
O
SPh
OBz
O
BzO
SPh
OBz
BzO
TMSOTf
TEP
OBz
O
O
OBz
BzO BzO
O
SPh
OBz
OAc
O
OAc
O
O
SPh
OTBDMS
O
O
O
O
OAc
O
SPh
OTBDMS
O
O
OH
O
SPh
OTBDMS
O
OAc
O
OAc
O
O
TBDMSO
O
O
O
O
TBDMSO
O
O
O OAc
O O
O
SPh
OTBDMS
HO
O
O
SPh
O
O
TBDMSO
O
O
O OAc
SPh
HO
O
O
O
O
O
O
TBDMSO
O
O
O OAc
O
SPh
O
Scheme 5
[TEP, triethylphosphite, is required to trap the transiently formed phenylsulphenyl ester which may activate the
acceptor resulting in the formation of a 1,6-anhydro derivative].
In the examples discussed above, only one type of anomeric leaving group has been used.
However, for the successful preparation of complex oligosaccharides often a range of different
leaving groups needs to be examined. An orthogonal glycosylation strategy uses two set of
chemically distinct (orthogonal) glycosyl donors activated under different conditions. In 1994,
Ogawa and co-workers proposed this strategy that reduces the manipulation at the
oligosaccharide stage.
In this approach two anomeric leaving groups (X and Y) are used acting either as anomeric
protecting group or as leaving group, depending on the activation conditions.
- 53 -
O
RO
O
X
O
O
O
RO
Y
O
RO
Disaccharide
Glycosyl acceptor
Glycosyl donor
O
O
Promotor-1
Y
HO
O
O
O
Promotor-2
X
HO
X
Glycosyl acceptor
Oligosaccharide
O
HO
Y
Promotor-1
O
O
O
Glycosyl acceptor
O
O
O
O
RO
Y
Oligosaccharide
Scheme 6
Example: Synthesis of chitotetraose oligosaccharide.
Kanie, O.; Ito, Y.; Ogawa, T. J. Am. Chem. Soc. 1994, 116, 12073.
OBn
HO
O
BnO
1
F
O
BnO
AcO
NIS
AgOTf
OBn
AcO
OBn
Nphth
Nphth
BnO
O
O
BnO
F
O
NPhth
4
OBn
SPh
Nphth
Cp2HfCl2
2 R = Ac
3R=H
AgClO4
HO BnO
3
OBn
AcO
OBn
O SPh
Nphth
OBn
Nphth
O
O
BnO
BnO
O
NPhth
O
O BnO
SPh
Nphth
OBn
5
NIS
AgOTf
OBn
AcO
O
BnO
HOBnO
OBn
Nphth
BnO
O
NPhth
O
OBn
O F
Nphth
O BnO
O
Nphth
O
Nphth
OBn
BnO
O
F
OBn
6
Scheme 7
4. - Chemoselective Glycosylation Reactions
This strategy uses the influence of the nature of the protecting groups on the reactivity of
donors and acceptors.
With respect to glycosyl donors, benzylated glycosyl donors (armed) are much more reactive
than acylated ones (disarmed). This difference makes chemoselective glycosylations possible,
- 54 -
the so-called Armed-Disarmed strategy. This strategy has been applied to several glycosyl
donors.
Armed-Disarmed strategy with NPGs
Benzylated pentenyl glycosides react faster than acylated ones.
OBn
BnO
OH
O
BnO
OPent
OBn
+
AcO
OBn
O
AcO
Armed
IDCP
OPent
OAc
Disarmed
BnO
O
BnO
O
OBn
AcO
O
OPent
OAc
AcO
OH
O
O
O
NIS/TfOH
O
OBn
BnO
O
BnO
O
O
OBn
AcO
O
AcO
O
O
OAc
O
O
O
O
Scheme 8
X
X
K1
O
O
O
O
OBn
O
OBn
O
Fast
O
OBn
OBn
X
X
K1
O
δ
Oδ
O
O
O
O
O
Slow
OBz
OBz
OBz
OBz
δ
Scheme 9
IDCP is appropriate for the coupling of some reactive (armed) NPGs but is not potent enough
for use with unreactive (disarmed) NPGs. For this purpose, NIS/Et3SiOTf or NIS/TfOH must be
employed.
In the cases where the nature of the protecting groups does not allow the application of the
armed disarmed strategy, two NPGs can still be coupled by use of an intermediate dibromination
step. Thus, depending on how the reaction is carried out, one can obtain either the glycosyl
bromide or a vicinal bromide.
Br
O
G
Br
Br2
O
R
Zn
Bu4NI
Br
O
G
O
G
R
O
O
G
Et4NBr
Br
O
R
Scheme 10
- 55 -
R
Br
O
O
G
R
Br
O
G
R
Br
Example, synthesis of Glycophosphatidylinositol Membrane-Bound Protein Anchors (GPI)
Roberts, C.; Madsen, R.; Fraser-Reid, B. J. Am. Chem. Soc. 1995, 117, 1546-1553.
Ph
OH
BnO
BnO
BnO
OH
BnO
O
BnO
Br2/Bu4NBr
BnO
O
AcO
BnOBnO
Br
Br
O
BnO
O
BnO
O
BnO
NIS/Et3SiOTf
O
BnO
O
O
AcO
Ph
O
O
O
O
Br
Br
O
BnO
O
OBn
BnO
OH
ClAcO
BnO
BnO
MPG
O
O
Br
Br
O
BnO
BnO
ClAcO
BnO
BnO
O
BnO
BnO
BnO
OBn
O
BnO
BnOBnO
NIS/Et3SiOTf
O
BnO
O
BnO
Br
Br
O
BnO
O
ClAcO
BnO
BnO
Zn/Bu4NI
O
BnO
O
OBn
BnO
BnO
BnO
O
BnO
O
GPI
O
O
BnO
O
O
O
Scheme 11
Armed-Disarmed strategy with thioglycosides
Protecting groups in the sugar ring and in the aglycone influence the reactivity of the donors.
Example: Veeneman, G. H.; van Boom, J. H. Tetrahedron Lett. 1990, 31, 275.
OBn
O
BnO BnO
SEt
OBn
armed
IDCP
+
HO BzO
OBz
O
91%
BnO BnO
disarmed
SEt
OBn
O
BnO
O
BzO
OBz
O
BnO BnO
armed
OBn
O
BnO
O BnO
1. NaOMe
2. NaH/BnBr/Bu4NI
OBz
OBz
disarmed
SEt
OBn
O
OBn
HO
OBz
O
BzO
disarmed
SEt
SEt
BnO BnO
OBz
IDCP
72%
OBn
O
BnO
O
OBn
O
BnO
BnO
O BzO
OBz
O
SEt
OBz
Scheme 12
The anomeric thio substituent also has an influence. Simple alkyl substituents such as
methyl, ethyl or isopropyl groups, show comparable reactivity towards thiophilic promoters.
However, a bulky alkyl substituent such as diciclohexylmethyl is much less reactive. This
allows the assembling of sugars in a chemoselective fashion.
- 56 -
Example: Boons, G. J.; Geurtsen, R.; Holmes, D. R. Tetrahedron Lett. 1995, 36, 6325.
OBn
O
BnO BnO
BnO BnO
OBn
O
SEt
OH
O
BnO BnO
+
OBn
BnO
O
IDCP
S
OBn
BnO BnO
O
S
OBn
Scheme 13
Phenylthio groups are less reactive than alkyl groups, but for chemoselective glycosylation,
the reactivity of aryl thioglycosides must be further adjusted by incorporation of electron
withdrawing or donating substituents.
It is important to point out that “armed” thioglycosides can be readily activated with
moderate iodonium sources such as IDCP or NIS. Activation of “disarmed” thioglycosides
requires the presence of a more powerful iodonium source. The combined use of NIS (1 eq) and
catalytic TfOH (0.015 eq) was shown to be particularly effective for this purpose.
Armed-Disarmed strategy with selenoglycosides
Van Boom demonstrated that alkylated phenylseleno glycosides can also be activated by the
thiophilic promoter IDCP to give O-glycosides in a similar way to thioglycosides.
However, fully benzoylated phenylseleno glycosides are not completely inert towards IDCP.
In some instances, orthoesters were detected.
So acylated phenylseleno glycosides can be considered as “pseudo disarmed” substrates. On
the other hand, performances of the same coupling in the presence of the powerful iodonium
source NIS-TfOH smoothly yield the β-linked disaccharide in 91% yield.
Example: Zuurmond, H. M.; Veeneman, G. H.; van der Marel, G. A. and van Boom, J. H.
Tetrahedron Lett. 1992, 33, 2063.
BnO
BnO
BzO
OBn
O
SePh
BnO
+
OBn
O
OH
BnO
IDCP
O
BnO
(87% α/β :4/1)
SePh
BzO
O
BzO
OBz
OBn
O
BzO
BzO
BnO
OBz
BnO
O
SePh
BzO
BzO
O
SePh
OBn
O
O
Ph
IDCP
60%
OBz
OBz
O
OH
SePh
BzO
OBz
BnO
O
O
SePh
BnO
OBn
BzO
BnO
OBz
+
O
SePh
BzO
OBz
BzO
OH
NIS-TfOH
O
OMe
BnO
91%
MeO
OBz
O
BzO
OBn
O
O
OBn
OBz
OBn
BnO
Scheme 14
- 57 -
Armed-Disarmed strategy with glycals
Finally, glycals can be also selectively activated by varying the protecting groups.
Example: Friesen, R. W.; Danishefsky, S. J. J. Am. Chem. Soc. 1989, 111, 6656.
BnO
BzO
HO
BzO
O
BnO
BnO
+
BnO
BnO
BnO
I
O
O
BzO
O
BnO
BzO
O
BzO
O
BzO
O
Pr3SnH
BzO
AIBN
O
O
O
O
O
O
O
O
O
OO
O
O
BnO
BnO
I
O
O
BnO
BnO
O
IDCP
OH
BzO
IDCP
BnO
I
O
O
O
O
O
Scheme 15
Tuning the glycosyl donor leaving group ability with a set of two groups, increases the
versatility of the armed–disarmed glycosylation strategy.
Chemoselective strategy with phenylseleno glycosides/thioglycosides
As expected, phenyl seleno glycosides are considerably more reactive than their thio
counterparts towards iodonium-ion mediated activation.
Example: Mehta, S.; Pinto, B. M. J. Org. Chem. 1993, 58, 3269-3276.
BnO
BnO
O
SePh
BnO
O
OH
OBn
+ BnO
OBn
IDCP
O
SEt
BnO
BnO
BnO
(79% α/β :3/1)
O
OBn
OBn
O
BnO
BnO
SEt
OBn
BzO
EtS
OBz
O
BzO
OBz
OH
+ BzO
SePh
BzO
O
SEt
NIS
TfOH
79%
OBz
BzO
OBz
OBn
O
O
BzO
BzO
O
BnO
OBn
Scheme 16
These results indicate that the intrinsic higher reactivity of phenyl selenoglycosides with
respect to the sulphur congeners significantly increases the scope of the armed-disarmed
strategy.
S. Ley and co-workers have developed a chemoselective strategy for oligosaccharide
assembly by tuning the reactivity of glycosyl donors with a set of two leaving groups and by
ester groups and spiroketals.
- 58 -
Donors and acceptors are grouped into four levels of reactivity (Fig. 2):
Oligosaccharide Assembly
Level 1
Most reactive
glycosyl donor
OBn
O
BnO
BnO BnO
FBnO
3
OBn
O
BnO BnO
R
R
O
O
MeO
2
4
SePh
MeO
R
R
O
O
MeO
5
MeO
R
R
O
O
OH
OBz
O
SePh
OBn
OBn
O
MeO
HO
Level 4
Not reactive
OAc
O
BnFO
FBnO
SePh
1
Level 3
change of Se to S
reduces reactivity
Level 2
electron-withdrawing groups
and/or fused rings reduce
donor reactivity
BnO
BnO
HO
OTPS
OBn
O
HO
SEt
7
O(CH2)8COOMe
9
SePh
MeO
OTPS
OH
O
R
R
O
O
MeO
SePh
OBz
OH
O
8
OTPS
OH
O
MeO
R
R
MeO
SEt
O
O
OMe
10
OBz
OH
O
SePh
6
MeO
R = Me, BDA R,R = -(CH2)4-, CDA
Fig.2
The general approach to the chemoselective synthesis of a trisaccharide by careful tuning of
glycosyl donor and glycosyl acceptor reactivity is outlined in Scheme 17.
OH
O
O
RO
RO
XR
High Reactive Donor
A
OH
O
RO
O
O
1 eq. NIS, cat. TfOH
(X = S or Se)
RO
XR
RO
XR
Acceptor and
Intermediate Reactive Donor
B
XR
Acceptor and
Low Reactive Donor
C
O
Intermediate
Reactive Donor
AB
1 eq. NIS, cat. TfOH
(X = S or Se)
O
RO
O
O
RO
O
O
RO
XR
ABC
Scheme 17
This methodology has been applied to the synthesis of high-mannose oligosaccharides.
- 59 -
Example: Grice, P.; Ley, S. V.; Pietruszka, J.; Osborn, H. M. I.; Priepke, H. W. M.;
Warriner, S. L. Chem. Eur. J. 1997, 3, 431-440.
BnO
OBn
O
BnO
BnO BnO
1
SePh
+
OBz
OH
O
MeO
O
O
MeO
BnO
NIS, cat.
TfOH
BnO BnO
MeO BzO
O
O
OO
MeO
O
O
BnO
OBz
OH
O
MeO
OBn
O
BnO BnO
MeO BzO
SEt
8
NIS, cat.
TfOH
1
+
MeO
O
O
MeO
5
BnO
SePh
OTPS
OH
O
NIS, cat.
TfOH
BnO BnO
MeO TPSO
O
O
MeO
OBn
O
BnO HO
OH
OBz
O
5 SEt
O
O
MeO
BnO BnO
MeO TPSO
O
O
MeO
SePh
HOHO
HO
HOHO
NIS, cat.
TfOH
OBn
O
Deprotection
HO O
MeO TPSO
O
O
O
O
MeO
O
BnO
14
OH
O
O
O
BnO
R
OBn
O
O
12, R = TPS
13, R = H
SEt
11
O
O
O
O
O
BnO BnO
SePh
MeO
MeO BzO
O
BnO
NIS, cat.
TfOH
HO
13+ 14
O(CH2)8COOMe
BnO
OBn
O
BnO
6
OBn
O
O
O
AgOTf, Br2
O
O
MeO
BnO BnO
O
O
MeO
MeO BzO
SePh
MeO
OTPS
OBn
O
BnO HO
O
O
SePh
6
OBn
O
BnO BnO
MeO BzO
O(CH
OBn
O
O
O
O
OBz
O
SEt
O
O
O
HO O
OO
OH
O
O
HO O
HO OH
HO OH
O
OH O
OH
O
O
OH OH
O
OH
HO
OH OH
OH
OH OH
OH
OH
O O(CH ) COOMe
2 4
High-mannose oligosaccharide
Scheme 18
5. One-pot multistep glycosylations
One-pot synthesis of oligosaccharides is often referred as a reactivity-based one-pot method
in which glycosyl donors with decreasing anomeric reactivities are allowed to react sequentially
in the same flask. This procedure, although is highly convenient because reduces the number of
steps considerably, has the inconvenience that the donor reactivities have to be carefully
adjusted which implies extensive protecting group manipulations.
Reactivity-based one-pot method
Tuning the reactivity of glycosyl donors by the influence of leaving and protecting groups,
together with the principle of orthogonal activation enabled a highly efficient tetrasaccharide
one-pot synthesis.
Example: Cheung, M.-K.; Douglas, N. L.; Berthold, H.; Ley, S. V.; Pannecoucke, X. M.
Synlett 1997, 257.
- 60 -
OAc
O
FBnO
FBnO
FBnO
F
(1.2 eq) 3
AgOTf
+
BnO
MeO
FBnO
OH
OBn
O
FBnO FBnO
CpHfCl2
4A MS
CH2Cl2
BnO
BnO
OAc
O
O
OBn
O
OBz
OH
O
OAc
O
FBnO
FBnO FBnO
SePh
BnO
NIS, cat.
TfOH
BnO
O
OBn
O
BnO
MeO BzO
SePh
SePh
(1.0 eq) 2
OO
MeO 6
(1.3 eq)
O
O
MeO
OAc
O
FBnO
FBnO FBnO
BnO
MeO
O
OBn
O
O
O
BnO
O
O
TPSO
MeOOMe
O
O
O
O
SePh
OTPS
OH
O
MeO
10
(1.6 eq)
O
MeO BzO
O
OMe
NIS, cat.
TfOH
O
O
OMe
MeO
overall yield, 21%
15
Scheme 19
Example: Synthesis of Cyclamycin 0
Raghavan, S.; Kahne, D. J. Am. Chem. Soc. 1993, 115, 1580-1581.
This is also a reactivity based one-pot procedure involving sulfinil glycosides. Groups in the
aglycon do the tuning of reactivity. This can be explained by taking into account that the
activation of sulfinil glycosides with Tf2O or TfOH begins with triflation of the sulfoxide.
O
O
S
O
S
F3 C
OY
rate limiting
O
CF3
S
O
O
S
R
+ YO
R
NO2 < H < OMe
R
Scheme 20
This step is rate limiting; therefore the reactivity of the glycosyl donor can be influenced by
manipulating the substituent in the para position of the phenyl ring in the order: NO2<H<OMe.
The reactivity difference between p-methoxyphenyl sulfinil donor and an unsubstituted phenyl
sulfinil glycosyl acceptor is large enough to permit selective activation. In addition, silyl ethers
are good glycosyl acceptors when catalytic TfOH is the activating agent because they react more
slowly than the corresponding alcohol. These features opened the way for one-pot synthesis of
- 61 -
Ciclamycin 0 trisaccharide in a stereoselective manner from the monosaccharide components in
one-step.
OH O
COOMe
O
S
O
OH
OH O
S
Me
OH O
Me
O
+
+
O
Me
O
OH
O
Me3SiO
1
Me
O
OH
O
Me
O
O
Me
+
OBn
HO
2
Me
Me
Me
O
OBn
O
O
Me
O
O
Ciclamycin 0
-70°
5
Me
HO
OBn
4
O
TfOH
OBn
+
-70°
O
fast
S
slow
Me
O
overall yield = 25%
3
O
O
TfOH
OBn
O
OBn
S
S
O
S
OMe
O
O
1
Scheme 21
The glycosylation takes place in a sequential manner, para-methoxyphenylsulfoxide 2 is
activated faster than phenyl sulphoxides 1, and 2 reacts preferentially with acceptor 3 using
triflic acid (TfOH) as promoter. In addition, while silyl ethers are stable to triflic anhydride
(Tf2O), they are good acceptors when the promoter is triflic acid; however, the HO-4 of 2 reacts
more slowly than the HO-4 of 3 because it has to be deprotected before reaction. In this way, the
reactivity of the reactants has been manipulated in order to obtain the trisaccharide in one-step.
Non-reactivity-based one-pot method
Recently Huang, Ye and co-workers have designed a general one-pot method independent of
differential glycosyl donors.
Example: Huang, X.; Huang, L.; Wang, H.; Ye, X.-S. Angew. Chem. Int. Ed. 2004, 43, 52215224.
The method is achieved by pre-activating the donor, that generates a reactive intermediate
that reacts with the acceptor that contains the same reactive leaving group. The process can be
repeated in the same vessel allowing the rapid assembly of oligosaccharides (Scheme 22).
- 62 -
RO
O
O
STol promoter
RO
O
STol
RO
HO
O
RO
O
O
promoter
STol
RO
X
reactive
intermediate
RO
O
O
RO
O
O
RO
STol
O
HO
RO
O
RO
O
O
O
STol
RO
X
reactive
intermediate
Scheme 22
The general conditions were established by using p-tolyl thioglycosides as building blocks,
and as the stoichiometric promoter, p-toluenesulfenyl triflate (p-TolSOTf) formed in situ from
p-toluenesulfenyl chloride (p-TolSCl) and AgOTf.
O
STol
RO
BnO
BnO
OAc
O
BnO
BnO
STol BnO
BnO
+ AgOTf
p-TolSCl
Acceptor
Product
BnO
OH
HO
O
O
STol BzO BzO
2
1
p-TolSCl (1 eq)
Et2O
2
STol AcO
O
AcO
OBz
3
4
3
p-TolSCl
BnO
HO
1
O
BnO
OAc
BnO
OAc
p-TolSCl
OAc
BnO
55% yield
~ 2 hours
O
2
O
BnO
4
O
O
BzO BzO
BzO
O
1 + AgOTf
5 min
-60ºC
15 min 15 min 5 min 5 min
RT
15 min 15 min 5 min 5 min
RT
-60ºC
-60ºC
15 min
3
AcO
-20ºC
4
O
AcO
Scheme 23
The tetrasaccharide Man-α(1,2)-Man-α(1,6)-Glc-α(1,6)-Glc was assembled in this way in
55% overall yield and in less that two hours.
6. Solid-phase oligosaccharide synthesis
The solid-phase synthesis SPS (also called SPOS: Solid-Phase Organic Synthesis) is a
methodology that performs the synthesis of a target compound on insoluble supports.
It offers several advantages over solution phase reactions:
•
Increased yields, because excess reagents can be used to drive the reaction to
completion.
•
Easy and simple purification processes, because removal of the by-products and
excess of reagents can be done by simply washing the resin.
- 63 -
OAc
OAc
•
Rapid overall process, purification of the reaction products is made at the end of the
synthesis minimizing the number of chromatographic steps required.
It is becoming a valuable alternative to traditional synthesis.
Bruce Merrifield was the chemist that in 1963, pioneered solid phase synthesis. For this
contribution, he earned the Nobel Prize of Chemistry in 1984.
The use of solid support for organic synthesis relies on three interconnected requirements:
Linker
Solid
Support
Functional
Group
Fig. 3
1. Solid support: A cross linked insoluble polymeric material that is inert to the conditions
of synthesis.
2. Linker: Some means of linking the functional group of the substrate to the solid phase
that permits selective cleavage of some or the entire product from the solid support
during synthesis to control the extent of the reaction, and finally, gives the desired
product.
3. Functional group: that requires a chemical protection/ deprotection strategy of the
reactive groups.
Merrifield developed a series of chemical reactions that were used to synthezise peptides
(Scheme 24). The carboxy terminal amino acid is anchored to a solid support. Then, the next
amino acid is coupled to the
Merrifield Peptide Synthesis on Solid Phase
first one. In order to prevent
NO2
Cbz N
H
CH3
O
PS
CH3
O
O
PS
CO2H
O
O
N
H
H
N
N
H
O
CH3
O
O
NO2
O
PS
Neutralization
CH3
NH2
O
N
H
H
N
Neutralization
Desattachment HO
Cbz
CH3
further chain growth at this
point,
the
amino
acid,
which is added, has its
Deprotection,
Cbz
H3C
CH3 O
H
N
H3C
2) Et 3N
H3C
H3C
N Cbz
H
CO2NHEt3
Coupling
Deprotection
1) HBr/AcOH
O
PS
H3C
DCC Cbz N
H
H3C
NO2
Cl Attachment
PS
CH3
O
1) HBr/AcOH
2) NaOH
CH3
CH3
O
N
H
the
CH3 O
H
N
O
N
H
L-leu-L-ala-gly-L-val
R. B. Merrifield, J. Am. Chem. Soc. 1963, 85, 2149
amino group blocked. After
Coupling
NH2
coupling
step,
the
protecting group is removed
CH3
CH3
from the primary amino
group and the coupling
reaction is repeated with the
Scheme 24
next amino acid. The process continues until the peptide or protein is completed. Then, the
molecule is cleaved from the solid support and any groups protecting amino acid side chains are
removed. Finally, the peptide or protein is purified to remove partial products and by-products.
- 64 -
Merrifield’s Solid Phase synthesis concept, first developed for the synthesis of peptides, has
also been extensively used for other biopolymers such as oligonucleotides.
Additionally, it has spread into every field where organic synthesis is involved. Many
laboratories and companies focus on the discovery of new chemistry (new reagents, new
reactions) suitable for SPS. It has contributed to a spectacular advance which profoundly
changed the approach for new drugs, new catalysts or new natural discovery.
Many laboratories and companies focused on the development of technologies such as
automated solid-phase synthesis. This has been set up for peptides and oligonucleotides
SPS of oligosaccharides simplify considerably the synthesis of such complex structures and
has had an immense impact on the chemistry and biochemistry of oligosaccharides. However, it
implies more problems than the SPS of peptides or oligonucleotides, because the preparation of
a specific carbohydrate requires the stereospecific formation of each new glycosidic bond in
high yield. Such processes have been demonstrated to be very sensitive even to slight structural
or electronic variations in the glycosyl donor or acceptor.
However, important progress in the field is currently taking place and this will provide an
important and fundamental impulse in the field of Glycobiology.
Central Aspects of Solid-Phase Oligosaccharide Synthesis. 12
Points to be considered:
1. The design of an overall synthetic strategy with either the 'reducing' or the 'nonreducing'
end of the growing carbohydrate chain attached to the support.
2. Selection of a polymer and linker which has to be inert to all reaction conditions during
the synthesis but has to be cleaved smoothly and effectively when desired.
3. A protecting-group strategy consistent with the complexity of the desired oligosaccharide
4. Stereospecific and high-yielding glycosylation reactions
5. 'On-bead' analytical tools that facilitate reaction monitoring and enable a rational
development of efficient protocols.
With regard the 1st point there are three synthetic strategies (Scheme 25):
12
Seeberger, P. H; Haase, W.-C. Chem. Rev. 2000, 100, 4349-4393
- 65 -
In the donor-bound strategy, the glycosyl donor is bound to the solid support by a suitable
hydroxyl group, and then reacted with solution phase acceptors. In the acceptor-bound strategy
the acceptor is attached to the solid support usually at the anomeric center. In the 3rd , strategy
acceptor or donor can be attached to the polymer and elongated differentially.
OR2
OR2
O
D
R 1O
A
HO
O
D
O
OP
OR1
R 1O
OR1
X
OH
X
OP
O
O
OR1
OR2
OR2
R 2O
A
A
OP
PO
OP
remove P
reiterate
OR2
OR2
O
O
O
HO
OR1
OR1
OR2
R2O
O
A/D
OR1
O
OR2
D
O
R2O
O
A/D
OP
OR1
O
remove P
reiterate
OR2
D
O
O
A/D
Y
OP
OR2
R2O
OR2
D
remove P
reiterate
OR1
OR2
R2O
Bi-directional strategy
O
OR1
OR1
O
O
A
R1O
OR1
R1O
Y
O
OP
OR2
D
O
D
OH
R2O
OR2
O
O
A
remove P
reiterate
OR1
OR2
R2O
O
OP
OR1
OR2
Acceptor-bound strategy
OR2
A
O
O
O
X
O
Donor-bound strategy
R 2O
OR2
O
R2O
OR1
Scheme 25
With regard to the 2nd point, there are different polymer and linker systems that are used in
SPS of oligosaccharides. Merrifield resin is a polystyrene resin that has been extensively used. It
has high loading capacity (1.2 mmol/g), requires swelling by the solvents for efficient reaction
to occur, it has low price, but is limited to solvents such as DMF, CH2Cl2, THF and dioxane.
Recent developments includes the grafting of polyoxoethylene onto polystyrene crosslinked
resins such as Tentagel and related resins. These have better swelling properties and are
compatible with water, but have lower loading properties (0.2-0.3 mmol/g) and higher price.
HO
HO
Cl
Cl
O
HO
Cl
O
O
n
O
O
n
O
n
Cl
Cl
Fig. 4
Cl
Merrifield 's resin
Tentagel
- 66 -
With regard to the linkers, they must fulfil the following requirements:
a) Must be inert to all reaction conditions
b) Determine protecting groups and coupling possibilities
c) Can be viewed as a protecting group
d) Orthogonal method for effectively cleavage under mild conditions.
Linker systems are:
i.
Silyl Ether Linkers
v.
Linkers cleaved by Hydrogenation
ii.
Acid- and Base-Labile Linkers
vi.
Photocleavable Linkers
iii.
Thioglycoside Linkers
vii.
Linkers
iv.
Linkers cleaved by Oxidation
cleaved
by
olefin
Metathesis.
With regard to the protecting groups, the most commonly used are:
Benzyl ethers, base-labile and acid-labile protecting groups, silyl ethers and allyl protecting
groups or others, specifically 4-azido-3-chlorobenzyl (ClAzb).
With regard to stereospecific and high-yielding glycosylation reactions, the gycosylating
agents used for SPS of oligosaccharides are:
i.
Glycosyl trichloroacetimidates
v.
Glycosyl Fluorides
ii.
Glycosyl sulfoxides
vi.
n-Pentenyl Glycosides
iii.
1,2-anhydrosugars
vii.
Glycosyl Phosphates
iv.
Thioglycosides
Finally, 'on-bead' analytical tools that facilitate reaction monitoring and enable a rational
development of efficient protocols.
These methods have had an immense impact on the development of solid-phase
oligosaccharide synthesis by allowing direct reaction monitoring. NMR and IR spectroscopy
together with MS spectrometry have been adapted for use on polymeric supports. These allow
on-bead characterization of oligosaccharides and their intermediates. The techniques used for
this purpose are:
A. HR-MS
B. High-Resolution Magic Angle Spining NMR
C. Gated Decoupling 13C-NMR
D. FT-IR Microspectroscopy
- 67 -
Pioneering Studies were carried out during the 1970s and 1980s.
Different strategies (donor- vs acceptor-bound synthesis), linkers, temporary protecting
groups and glycosylating agents were explored.
Example: Synthesis of α-(1→6)-trisaccharide.
Fréchet, J. M. J.; Schuerch, C. J. Am. Chem. Soc. 1971, 93, 492-496.
This strategy was quite successful in the preparation of α-linked 1→6-oligomers.
Drawbacks: long reaction times and the failure to selectively synthesize β-linked glycosides.
O
NO2
O
BnO
O
O
BnO
Br
HO
BnO
2
O
BnO
BnO
O
2,6-lutidine, 2 d, 65 ºC, 96%
1
NO2
O
OBn
3
OH
MeONa/MeOH
BnO
reiterative coupling/
deprotection
O
BnO
BnO
O
quant.
90%
4
O
O
NO2
O
BnO
O
BnO
O
BnO
BnO
O
2) SMe2, -78 ºC
79-95%
O
BnO
BnO
1) O3, -78 ºC
51-91%
BnO
O
BnO
BnO
BnO
O
O
BnO
O
BnO
BnO
O
BnO
NO2
O
BnO
BnO
O
O
BnO
BnO
O
6
5
OH
: Merrifield's resin
Scheme 26
Example: Synthesis of a chitobiose derivative.
Excoffier, G.; Gagnaire, D.; Utille, J.-P.; Vignon, M. Tetrahedron 1975, 31, 549-553
OH
O
O
1) pyridine, 7 d
O
O
HO
O
Ph
O
AcHN
OBn
21
2) PhCOCl, pyridine
3) hydrazinium acetate,
pyridine, AcOH, 50 ºC
Cl
20
OAc
AcHN
Cl
23
O
OAc
AcO
AcO
Hg(CN)2
85%
1) NaOMe, MeOH
2) Ac2O, pyridine
O
O
O BzO
O
AcHN
AcHN
OBn
24
OAc
AcO
AcO
OAc
O
O BzO
O
AcHN
AcHN
OBn
25
51%, based on 22
: "popcorn" polystyrene
- 68 -
O
AcHN
OBn
22
O
AcO
AcO
Scheme 27
O
BzO
HO
Drawback of "popcorn" polystyrene: partial solubility and thus, considerable loss of
material during the synthesis, reduced overall yield.
Major advances (1990s up to now) in solid-phase oligosaccharide synthesis includes:
1. Development of more powerful glycosylating agents of improved selectivity.
2. Greater diversity of available protecting groups.
3. New analytical techniques.
4. Automatization.
This opens the window for the rapid future development which was briefly glanced at by the
pioneers.
Examples:
A. Donor-Bound Glycosylation Strategy
a) Danishefsky, S. J.; Bilodeau, M. T. Angew. Chem. Int. Ed. Engl. 1996, 35, 1380-1419.
b) Seeberger, P. H.; Bilodeau, M. T.; Danishefsky, S. J. Aldrichimica Acta 1997, 30, 75-92
1,2-anhydroglycal method.
O
SiPh2
O
O
DMDO
O
O
O
O
O
O
O
52
O
50
O
SiPh2
O
OH
O
SiPh2
O
O
O
O
O
O
ZnCl2
53
SiPh2
O
OH
O
O
O
O OH
O
O
O
OH
O
TBAF, AcOH
O
O
O
OH
O
O
O
O
O
O
OH
O
O
O
O
O
O
O
O
OH
54
O
O
51
O
O
OH O
O
OH
BnO
BnO
O
55
32% overall
BnO
BnO
O
: Merrifield's resin
Scheme 28
Drawback of the donor-bound strategy:
Most side reactions during glycosylations involve the glycosyl donor. Any side reaction in
the donor attached to the resin will provoke termination of chain elongation. The consequence is
a reduction of the overall yield.
However, an impressive array of complex oligosaccharides has been synthesized by
Danishefsky and co-workers using the glycal assembly method under this strategy.
- 69 -
B. Acceptor-Bound Glycosylation Strategy
Example.: Wang, Z.-G.; Douglas, S. P.; Krepinsky, J. J. Tetrahedron Lett. 1996, 39, 6985-6988.
Trichloroacetimidate method.
HODOX-PEGM,
DBBOTf, 4 Å MS,
-45 ºC
OBn
O
AcO
BnO
O
CCl3
PhthN
OBn
NH
BnO
OBn
OBn
HO
BnO
MeOH
57
OAc
O
BnO
BnO
O
ODOX-PEGM
PhthN
56
DBU
O
AcO
BnO
O
O
BnO
ODOX-PEGM
O
PhthN
59
CCl3
NH
PhthN
DBBOTf, 4 Å MS, -45 ºC, 95%
58
BnO
OAc
O
BnO
BnO
OBn
OBn
O
O
BnO
O
PhthN
O
ODOX-PEGM
BnO
PhthN
60
Scheme 29
Excess of donors are used and the overall yields are good and side products are washed away
after each coupling. For this reason, the acceptor-bound approach has generated an immense
interest in the solid-phase oligosaccharide synthesis.
C. Bidirectional Strategy
Elongation of the growing oligosaccharide in both directions requires two sets of orthogonal
glycosyl donors. Examples:
a) Ito, Y.; Kanie, O.; Ogawa, Y. Angew. Chem., Int. Ed. Engl. 1996, 35, 2510-2512.
b) Kanie, O.; Ito, Y.; Ogawa, T. J. Am. Chem. Soc. 1994, 116, 12073-12074.
O
O
OH
O
O
NH2
O
O
O
HO
BnO
NH
62
O
SEt
O
O
PyBOP, DIPEA
O
HO
BnO
BnO
O
SEt
BnO
61
63
OBn
O
BnO
BnO
BnO
O
64
CCl3
NH
TMSOTf, 4Å MS
OBn
BnO
BnO
O
BnO
O
O
O
BnO
SEt
BnO
65
O
OH
O
O
OBn
O O
66
NIS/TMSOTf, 4Å MS
BnO
BnO
O
O
O
BnO
BnO
67
O
BnO
O
O
O
O
O
O
60% overall
: TentaGel
Scheme 30
- 70 -
In the reaction scheme, first the acceptor containing a potential leaving group is bound to the
resin. Reaction with the donor is performed under different conditions. Then an acceptor is
made to react with the initial anomeric leaving group
Automated Solid-Phase Synthesis
P. Seeberger and co-workers have demonstrated that relatively simple carbohydrates can be
prepared on a machine that executes a coupling cycle, including steps for glycosylation and
deprotection.
The first automated solid-phase oligosaccharide synthesizer was used to prepare structures as
large as branched dodecamers within less than one day. A re-engineered peptide synthesizer
containing a coolable reaction vessel was used. As linker they used octenediol that can be
attached to the resin through either ester or ether linkage. Each monosaccharide has a protection
group pattern that permits the selective deprotection of a single hydroxyl group. As donors
glycosyl phosphates were used that are readily obtained by reaction with diphenylphosporyl
chloride following Sabesan’s method13. These donors are activated with a Lewis acid such as
TMSOTf and have reactivity similar to trichloracetimidates.
RO
OPh
OPh
DMAP, CH2Cl2
Cl
O
OH
O
P
O
RO
O
O P OPh
OPh
R'OH, TMSOTf
O
RO
OR'
MeCN, -78º
Scheme 31
The automated synthesis starts with glycosylation of a resin-bound acceptor producing a
coupling product that may be subsequently deprotected. Iteration of coupling and deprotection
cycles with phosphate donors followed by cleavage of the resin-bound oligosaccharides and
purification gives the products.
Fig. 5
Products
Example: The Synthesis of Protected Tumor-Associated Antigen and Blood Group
Determinant Oligosaccharides
13
Sabesan, N.; Neira, S. Carbohydr. Res. 1992, 223, 6453
- 71 -
Routenberg K. L., Seeberger, P. H. Angew. Chem. Int. Ed, 2004, 43, 602-605
The Lewis blood group oligosaccharides are a family of fucosylated, ceramide-containing
glycoesphingolipids decorating the exterior of healthy and disease-derived cells.
Lewis type penta- and hexasaccharides are part of the inflammatory cascade and have been
implicated in bacterial and viral infection as well as in autoimmune diseases. The biological
importance of the Lewis antigens has made them targets of intense examination.
Lewis X
Lewis Y
Lewis Y-Lewis X
monosaccharide building blocks 4-8.
Bn=benzyl, Bu=butyl, Fmoc=9-fluorenylmethoxycarbonyl, Lev=levulinoyl, Piv=pivaloyl,
TCA=trichloroacetamide
Scheme 32
Fmoc carbamate and levulinoyl ester were selected as temporary protecting groups because
both of them are completely orthogonal and are easily removed with piperidine and hydrazine,
respectively. As linker it was used octenediol that, in this case, reacted with carboxy-terminated
polystyrene resin resulting in an ester linkage, which was rapidly cleaved with a strong base at
the end of the synthesis. Glycosyl phosphates were used as donors.
Initial glycosylation of resin-bound acceptor 9 produces a coupling product that may be
subsequently deprotected. Iteration of coupling and deprotection cycles with phosphate donors
4-8 followed by cleavage of the resin-bound oligosaccharides and purification gives 1-3.
The automated synthesis of pentasaccharide 1, hexasaccharide 2, and nonasaccharide 3 on
the 25-mmol scale, is represented in Scheme 32. Each coupling is promoted with TMSOTf, in a
ratio 1:1 with the donor and is repeated 2 or 3 times. Washing with piperidine or with hydrazine
liberates the appropriate hydroxyl group. Finally treating with an excess of NaMeO/MeOH
several times, liberates the oligosaccharide from the resin.
- 72 -
Scheme 33
Example: Synthesis of a dodecasacharide
Plante, O. J. ; Palmacci, E. R.; Seeberger, P. H. Science 2001, 291, 1523; Bartolozzi, A.
Seeberger, P. H., Current Opinion in Structural Biology 2001, 11, 587.
Scheme 34
- 73 -
Each cycle involved the delivery and coupling of a building block to a growing, polymerbound oligosaccharide chain and the removal of a protecting group to expose a unique hydroxyl
group for attachment of the next carbohydrate. Stepwise coupling yields, greater than 94%, were
obtained in the assembly of linear and branched carbohydrates.
Finally, very recently P. Seeberger and co-workers have reported the use of a Microreactor
based method for performing glycosylation reactions very rapidly over a wide range of reaction
conditions.
Ratner, D. M.; Murphy, E. R.; Jhunjhunwala, M.; Snyder, D. A.; Jensen, K. F.; Seeberger, P.
H. J. Chem. Soc., Chem. Comm. 2005, 578-580.
The Silicon microfluidic microreactor (Fig. 6) was designed with three primary inlets to mix
and react glycosylating agents, acceptor and promoter. Once mixed the reactants, they enter a
reaction zone which is terminated by a secondary inlet used to quench the reaction, and after
that, the quenched reaction stream exits the reactor for collection and analysis.
Fig. 6
This method of optimization is currently under development and, together with
automatization, will probably have a tremendous impact on the progress of Glycochemistry.
=====================================
- 74 -
75