Synthesis of two tetra- and four pentasaccharide fragments of Shigella flexneri serotypes 3a and X O-antigens from a common tetrasaccharide intermediate

FULL PAPER

DOI: 10.1002/ejoc.200800693

Synthesis of Two Tetra- and Four Pentasaccharide Fragments of Shigella

flexneri Serotypes 3a and X O-Antigens from a Common Tetrasaccharide

Intermediate

Julien Boutet^[a,b]and Laurence A. Mulard*^[a]

Keywords: Carbohydrates / Glycosylation / Antigens / Solvent effects / Protecting groups

Relying on trichloroacetimidate chemistry, six tetra- and

pentasaccharide fragments of the {2)-[α- -Glcp-(1Ǟ3)]-α-

-Rhap-(1Ǟ2)-α- -Rhap-(1Ǟ3)-[AcǞ2]-α- -Rhap-(1Ǟ3)-β-

with a D acceptor and subsequent partial or total deprotec-

tion, respectively. Additionally, the selective removal of the

2_A-levulinoyl protecting group in 9 allowed for chain elong-

ation at this position. The glycosylation of the resulting ac-

ceptor with a D donor, and subsequent partial or total depro-

tection, gave branched pentasaccharides D(E)AB_AcC and

D(E)ABC. All targets are parts of the O-antigen of Shigella

flexneri 3a, a prevalent serotype. Non-O-acetylated oligosac-

charides are shared by the S. flexneri serotype X O-antigen.

Germany, 2008)

D

L

D-

GlcpNAc-(1Ǟ}_n((E)AB_AcCD)_npolymer were synthesized as

their propyl glycosides by use of a common fully protected

(E)AB_AcC intermediate (9). Tetrasaccharide 9 derived from

the condensation of an EA donor and a B_AcC acceptor. Partial

and full deprotection gave free tetrasaccharides (E)AB_AcC

and (E)ABC, respectively. Alternatively, 9 was converted into

a trichloroacetimidate donor, which provided linear penta-

saccharides (E)AB_AcCD and (E)ABCD, following a reaction

types.^[7]Several serotypes are isolated from individual

patients, emphasizing the need for a multivalent vaccine

providing broad serotype coverage. Although strain preva-

lence highly depends on geographic areas, serotype 2a and,

among others, serotype 3a are predominant.^[5,6]Investi-

gations in the field indicated that clinical infection protects

against subsequent exposure to a homologous serotype.^[8,9]

This is a good indication that in addition to playing a key

role in bacterial virulence and resistance to innate immu-

nity,^[10]S. flexneri O-Ags are also crucial targets of the host

protective adaptive immunity. All S. flexneri serotypes but

one share a linear tetrasaccharide backbone, made of 3 α-

linked -rhamnosyl residues (A, B, C) and a 2-acetamido-

2-deoxy-β--glucopyranosyl residue (D). Tetrasaccharide

ABCD (I) is the basic repeating unit of serotype Y (Fig-

ure 1). Additional serotype specificity is associated with the

presence of branched α--glucopyranosyl (E) and O-acetyl

(O-Ac) decorations (Figure 1).^[7]It is noteworthy that LPS

glucosylation affects the S. flexneri 5a O-Ag conformation,

resulting in a shortened length.^[11]A strong impact on bac-

terial virulence was hypothesized.^[10]However, the influence

of O-acetylation remains unknown.

Introduction

Enteric infections, which are responsible for some 1.7–

2.5 million deaths per year, rank third among all causes of

disease burden worldwide.^[1]Most of these deaths occur in

resource-poor countries, particularly in children younger

than five years old. In this population, enteric infections

are the second most common cause of death (18%) after

pneumonia (19%).^[2]A major drawback to the efficient pre-

vention of diarrhoeal diseases through vaccination is the

large number of pathogens involved. Among those is Shig-

ella, a Gram-negative enterobacterium causing shigellosis

or bacillary dysentery, which is an invasive infection of the

human colon often associated with blood and mucus in the

stool. Humans are the only reservoir for this highly con-

tagious infection, which is also increasingly associated with

antibiotic resistance.^[3,4]In the absence of a vaccine, shigel-

losis remains a major health concern.^[5]Shigella flexneri,

one of the four Shigella species, is the one most frequently

isolated worldwide. It is endemic to developing countries

and prevails in children below five years old.^[5,6]Based on

the carbohydrate repeating unit of its O-antigen (O-Ag),

which is the specific polysaccharide part of the bacterial

lipopolysaccharide (LPS), S. flexneri is divided into 15 sero-

In order to gain a better understanding of the key role

played by S. flexneri O-Ag decorations in bacterial viru-

lence and resistance to innate immunity by investigating

conformational behaviour, we initiated our study on S. flex-

neri 5a^[11,12]and S. flexneri 2a.^[13,14]More recently, we inves-

tigated additional serotype-specific glucosylation and O-

acetylation patterns.^[15]In a continuation of that study, we

focused herein on S. flexneri 3a. The successful use of well-

[a] Institut Pasteur, Unité de Chimie des Biomolécules, CNRS

URA 2128,

28 rue du Dr Roux, 75015 Paris, France

Fax: +33-1-40613820

E-mail: laurence.mulard@pasteur.fr

[b] Université Paris Descartes, 4 rue de l’observatoire,

75006 Paris, France

5526

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Shigella flexneri Serotypes 3a and X O-Antigens

Figure 1. Repeating units of the O-Ags of S. flexneri serotypes Y (I), X (II) and 3a (III).

defined synthetic fragments of the native polymer to probe access to tetrasaccharides 1 and 2, respectively. Alterna-

O-Ag structure and antibody recognition was demonstrated tively, the selective removal of the levulinate at position 2_A

by others for serotype Y^[16]and by us for serotype 5a^[11]of tetrasaccharide 9 would result in acceptor 10, whose con-

and 2a.^[14]In this context, we report below on the synthesis densation with the known glucosamine donor 11^[23,24]

of fragments of S. flexneri serotype X and 3a O-Ags, would give a precursor to both pentasaccharides 3 and 4.

whose repeating units are the branched pentasaccharides Furthermore, the selective cleavage of the allyl aglycon and

(E)ABCD (II) and (E)AB_AcCD (III),^[17]respectively (Fig- subsequent trichloroacetimidate activation would turn 9

ure 1). We note that the S. flexneri X O-Ag is the non-O- into donor 12. The latter should react with known acceptor

acetylated form of the S. flexneri 3a O-Ag.

13^[24,25]to give a pentasaccharide serving as a key interme-

diate for both mono-acetate 5 and fully deprotected 6.

Results and Discussion

Following our recent contribution^[15]on the synthesis of

fragments EA, D(E)A, CD(E)A, _AcCD(E)A, BCD(E)A and

Synthesis of the EA Disaccharide Donor 7

B_AcCD(E)A, we report herein on the chemical synthesis of

The E–A linkage is the only 1,2-cis glycosidic linkage in

tetrasaccharides (E)AB_AcC (1) and (E)ABC (2), as well as the targeted sequences, and for that reason, it was built at

on that of pentasaccharides D(E)AB_AcC (3), D(E)ABC (4), an early stage in the synthesis. The known disaccharide

(E)AB_AcCD (5), and (E)ABCD (6). Contrary to previous 14,^[26]prepared as described,^[15]served as an exquisite pre-

work on S. flexneri Y, ^[18]2a,^[19]and 5a^[20]where methyl gly- cursor to 7 (Scheme 2). However, levulinoylation at position

cosides were the targets, all fragments bearing either - 2_Aof 14 could not be completed and resulted in an insepa-

rhamnoside C or 2-acetamido-2-deoxy--glucopyranoside rable 3:2 mixture of levulinate 15 and starting 14 when at-

D at their reducing end were synthesized as their propyl tempted by the treatment of 14 with levulinic acid, DCC,

glycosides according to the following reasoning. All the gly- and DMAP in CH₂Cl₂, according to a known protocol.^[27]

cosylation steps relied on highly efficient trichloroacetimid- Analogously to previous observations,^[12,26]the failure to go

ate (TCA) chemistry,^[21]thus requiring temporary masking to completion was tentatively explained by the 2,3,4,5-tetra-

at the anomeric position of the various building blocks. Al- O-benzyl-α--glucopyranosyl residue at O-3_Acausing steric

lyl glycosides match this requirement and are often ef- hindrance at OH-2_Ain 14. Benzoylation at OH-2_Asuc-

ficiently used as intermediates in complex oligosaccharide ceeded when performed at 70 °C.^[26]However, the conver-

synthesis.^[22]Interestingly, in addition to being orthogonal sion of 14 to 15 with levulinic acid reached 80% at best

to a number of protecting groups, allyl ethers are easily re- when heating the reaction mixture or changing the solvent.

duced to propyl ethers upon hydrogenation with Pd/C as Therefore, other reagents were envisioned. Instead of levuli-

catalyst.^[15]We took advantage of this property to block noyl chloride, which is known to form labile pseudo es-

the reducing end of the oligosaccharide targets in a form ters,^[28]we investigated the use of levulinic anhydride.^[29,30]

mimicking the anomeric group found in the O-Ag.

Under conventional conditions, the reaction was sluggish.

The combined retrosynthetic analysis of 1–6 led to two However, increasing the amount of levulinic anhydride and

major conclusions, which served as a basis for the whole DMAP up to 10 and 5 equiv., respectively, while performing

strategy (Scheme 1): (i) fully protected tetrasaccharide the esterification at 50 °C allowed for complete conversion.

(E)AB_AcC (9), bearing a levulinoyl group at position 2_Aand The fully protected disaccharide 15 was isolated in an excel-

an allyl aglycon, could serve as a common intermediate to lent 95% yield. Allyl glycoside 15 was then converted to

all desired targets and (ii) the levulinoyl group, being or- hemiacetal 16 (92%) following a two-step selective deal-

thogonal to an acetyl group, could serve as a suitable pro- lylation procedure involving: (i) the isomerisation of the al-

tecting group, ensuring anchimeric assistance when using lyl ether into the corresponding prop-1-enyl ether with a

donors activated at position 1_Aor 1_B. Based on these as- cationic iridium complex^[31]and (ii) subsequent iodine-me-

sumptions, tetrasaccharide 9 would result from the conden- diated hydrolysis.^[32]Finally, the EA disaccharide donor 7

sation of an EA donor (7) and a BC acceptor (8), both of was obtained as an anomeric mixture in 97% yield by re-

which are readily available from commercial monosaccha- acting hemiacetal 16 with trichloroacetonitrile with a cata-

rides. Partial and full deprotection of 9 should provide easy lytic amount of DBU.

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J. Boutet, L. A. Mulard

FULL PAPER

Scheme 1. Retrosynthetic analysis of propyl glycosides 1–6.

fication. Alternatively, taking advantage of the easy accessi-

bility of large amounts of crystalline diol 18,^[15]a key pre-

cursor to donor 7, alcohol 26 was prepared in 88% yield

by the regioselective benzylation at position 3 of 18 upon

treatment of stannylene intermediate 19 with benzyl bro-

mide and caesium fluoride.^[36]Additionally, our investiga-

tion on allyl orthoester 22, obtained in 5 steps and 76%

yield from -rhamnose through diacetate 20 and diol 21,^[37]

opened new routes to alcohol 26. On one hand, the

TMSOTf-mediated intramolecular rearrangement of 22

gave acetate 23 (88%), which was turned into 26 upon treat-

ment with methanolic MeONa (92%), in a strategy similar

to one involving the corresponding methyl orthoester 17.

Scheme 2. Synthesis of the EA disaccharide donor 7: (a) Levulinic

anhydride, DMAP, pyridine, 50 °C, 95%; (b) i; catalytic

–

[Ir(COD){PCH₃(C₆H₅)₂}₂]⁺PF₆, THF, room temp.; ii. I₂, THF/

H₂O, room temp., 92%; (c) CCl₃CN, DBU, DCE, –5 °C, 97 %.

On the other hand, heating the 3:2 mixture of mono-O-

Synthesis of the BC Disaccharide Acceptor 8

1

acetylated regioisomers 24 (δ_C-1= 92.4 ppm, J_C,H

=

170.9 Hz and δ_H-1= 5.15 ppm, δ_H-2= 5.41 ppm), and 25

(δ_C-1= 93.32 ppm, J_C,H= 158.7 Hz and δ_H-1= 5.67 ppm,

δ_H-2= 4.16 ppm), resulting from the 10% aqueous HCl-

mediated hydrolysis of orthoester 22, in allylic alcohol con-

taining acetyl chloride gave 26 in 85% yield from the diben-

zyl orthoester. This alternative conversion of 22 into 26 is

Disaccharide 8 could be most conveniently prepared by

the condensation of known acceptor^[33]30 and the as yet

undisclosed trichloroacetimidate 29. The latter was ob-

tained from alcohol 26, which often derives from the high-

yielding BF₃·OEt₂-^[34]or TMSOTf-mediated^[35]allyl glycos-

idation of methyl orthoester 17 and subsequent transesteri-

1

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Shigella flexneri Serotypes 3a and X O-Antigens

Scheme 3. Synthesis of the B monosaccharide donor 29: (a) see ref.^[34](b) see ref.^[35](c) i. Bu₂SnO, 4-Å MS, toluene; ii. CsF, BnBr, DMF,

88%; (d) K₂CO₃, MeOH; (e) NaH, BnBr, DMF, 76%; (f) TMSOTf, 4-Å MS, CH₂Cl₂, 88%; (g) NaOMe, MeOH, 92%; (h) 10% aqueous

–

HCl, EtOAc; (i) AllOH, AcCl, 85%; (j) Levulinic acid, DCC, DMAP, room temp., 89%; (k) i. catalytic [Ir(COD){PCH₃(C₆H₅)₂}₂]⁺PF₆,

THF, room temp.; ii. I₂, THF/H₂O, room temp., 96%; (l) CCl₃CN, DBU, DCE, –5 °C, 92 %.

comparable to more conventional ones.^[34,35]According to performed on 7 g of 18. The removal of the 2_B-levulinoyl

this route, alcohol 26 was isolated in 72% from -rhamnose ester of 31 by reaction with hydrazine in pyridine/AcOH

after 6 steps with only one chromatographic purification. gave the required disaccharide acceptor 8 (81%, Scheme 4).

Despite a larger number of steps, we found this route to be

superior to that involving diol 18 (68% from -rhamnose)

since the overall yield was comparable, but tin was avoided

(Scheme 3).

Synthesis of the (E)AB_AcC and (E)ABC Tetrasaccharides 1

and 2

Alcohol 26 was treated with levulinic acid in the presence

of DCC and DMAP to give ester 27 (89%), which was con-

verted to hemiacetal 28 (96%) upon anomeric deallylation.

The latter was then activated as the trichloroacetimidate 29

(92%). The TMSOTf-mediated condensation of donor 29

and acceptor 30,^[38]resulting from the regioselective acety-

lation of diol 18, was attempted in both CH₂Cl₂and tolu-

ene. In toluene, the fully protected disaccharide 31 was iso-

lated in higher yield, reaching 89% when the reaction was

Performing the condensation of donor 7 and acceptor 8

in CH₂Cl₂containing a catalytic amount of TMSOTf gave

the fully protected tetrasaccharide 9 in 69% yield. In tolu-

ene, the yield of 9 reached 75% and was even brought up

to 92% when the condensation was performed on 4 g of 8

instead of 200 mg (Scheme 5). The NMR analysis of 9,

1

showing J_C,H= 176.6 Hz, indicated an α-AB linkage and

confirmed that the levulinic ester had played its role as a

participating group. The selective removal of the levulinoyl

group gave alcohol 10 (89%), which was submitted to hy-

drogenolysis and concomitant Pd/C-mediated allyl re-

duction under neutral conditions to give the mono-O-acety-

lated tetrasaccharide 1 (76%). Alternatively, refluxing 9 in

methanolic MeONa gave diol 32 (95%), which was then

converted into the target tetrasaccharide 2 (81%).

Synthesis of the D(E)AB_AcC and D(E)ABC

Pentasaccharides 3 and 4

Alcohol 10 served as an ideal intermediate in the synthe-

sis of pentasaccharides 3 and 4 (Scheme 6). As for the prep-

aration of pentasaccharide BCD(E)A,^[15]we reasoned that

a controlled transesterification would allow for the selective

Scheme 4. Synthesis of the BC disaccharide acceptor 8: (a) i. MeC-

(OMe)₃, PTSA, CH₃CN; ii. 80% aqueous AcOH, quant.; (b) cata-

lytic TMSOTf, 4-Å MS, –78 °C, toluene, 89%; (c) hydrazine hy-

drate, pyridine/AcOH (3:2, v/v), room temp., 81%.

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J. Boutet, L. A. Mulard

FULL PAPER

Scheme 5. Synthesis of tetrasaccharides 1 and 2: (a) catalytic TMSOTf, 4-Å MS, –78 °C, toluene, 92%; (b) hydrazine hydrate, pyridine/

AcOH (3:2, v/v), room temp., 89%; (c) NaOMe, MeOH, reflux, 95%; (d) Pd/C, H₂, EtOH, 76% 1, 81% 2.

removal of vicinal acetates in the presence of an isolated 0.5 equiv. had an additional benefit as seen from Entries 2

2_C-acetate. Therefore, the triacetate donor 11, having an N- and 3 of Table 1. In addition to the observed significant

trichloroacetyl (Cl₃Ac) protecting pattern, was selected as a effect of the reaction temperature (Table 1, Entries 3–5), in-

precursor to residue D. When tetrasaccharide 10 and donor creasing the amount of donor 11 from 1.3 to 2 equiv. had

11 were reacted in CH₂Cl₂containing TMSOTf under con- the most visible impact (Table 1, Entries 4, 7, and 8). Tak-

ditions successfully used for the condensation of disaccha- ing into account all of these observations, pentasaccharide

ride 14 and 11,^[15,24]the condensation product 34 was iso- 34 was isolated in a satisfactory 82% yield (Table 1, Entry

lated in only 20% yield, together with unreacted oxazoline 8). Conversion of 34 to triol 35 was the next step. Contrary

33 (40%, δ_H-1= 6.35 ppm, J_1,2= 7.5 Hz).^[23]This result was to previous observations on a related system,^[15]methanolic

tentatively explained by the increased steric hindrance at K₂CO₃was poorly selective. The regioselectivity was barely

the 2_A-hydroxy group on going from 14 to 10. Based on improved with Et₃N/MeOH/H₂O or methanolic MeONa

this hypothesis, attempts to improve the outcome of the gly- (data not shown). Under the best identified conditions, triol

cosylation relied on investigating the impact of four param- 35 and tetraol 36 were isolated following column

eters, including the solvent, temperature, and the amounts chromatography in 56 and 30% yield, respectively. The

of donor 11 and TMSOTf (Table 1). Comparison of Entries high-pressure Pd/C-mediated hydrogenation^[15]of 35 and

1 and 6 with 2 and 7 of Table 1, respectively, shows that for 36, allowing concomitant benzyl hydrogenolysis, allyl re-

this specific condensation, toluene is a better solvent than duction, and trichloroacetamide reductive hydrodechlorina-

CH₂Cl₂, although the improvement in yield was minor. tion, gave the pentasaccharide targets 3 (74%, 2 d) and 4

However, increasing the amount of TMSOTf from 0.3 to (74%, 1 d), respectively. The transformation of tri-

Scheme 6. Synthesis of pentasaccharides 3 and 4: (a) TMSOTf (0.5 equiv.), 4-Å MS, –40 °C, toluene, 82%; (b) NaOMe, MeOH, room

temp., 25 min, 86%; (c) Pd/C, H₂, 45 bar, EtOH, 74% 3, 74% 4.

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Shigella flexneri Serotypes 3a and X O-Antigens

chloroacetamide 35 into the 2_C-acetate target 3 was much The anomeric deallylation of 9 gave hemiacetal 37 (90%),

faster than the analogous conversion performed to obtain which was converted into trichloroacetimidate 12 (88%) fol-

_AcCD(E)A (76%, 10 d) and B_AcCD(E)A (70%, 10 d).^[15]lowing a reaction with trichloroacetonitrile. Donor 12 was

Interestingly, monitoring the conversion by mass and NMR subsequently reacted with acceptor 13.^[24,25]As an alterna-

spectroscopy in the latter cases showed that the chloroacet- tive to its known synthesis,^[25]the latter was obtained in two

amide to acetamide conversion was the rate-limiting step. steps from the known 4,6-O-isopropylidene acetal do-

Steric hindrance at NH-2_Dwas hypothesized.^[15]The data nor^[15,24]38. Trichloroacetimidate 38 was treated with allyl

presented here for obtaining pentasaccharide D(E)ABC (3), alcohol and a catalytic amount of TMSOTf in CH₂Cl₂to

with monosaccharide D acting as an end-chain residue sup- give allyl glycoside 39 (82%). The saponification of 39 and

port this hypothesis.

subsequent N-acetylation of the resulting amino alcohol

gave 13 (71%). The attempted condensation of 12 and 13

was performed under conditions that took advantage of the

knowledge gained in the synthesis of S. flexneri 3a oligosac-

charides as well as of the expertise derived from studying a

related system in the S. flexneri 2a series.^[22]Thus, tetrasac-

charide 12 was treated with acceptor 13 (2.5 equiv.) and

TfOH (0.9 equiv.) in toluene at 70 °C to give the condensa-

tion product 40 in a satisfactory 78% yield. The use of a

large excess of acceptor 13 was found necessary to over-

come its low reactivity, which leads to the hydrolysis of un-

reacted 12.

Table 1. Coupling of monosaccharide donor 11 and tetrasaccharide

acceptor 10 under various conditions.

Entry

TMSOTf

[equiv.]

Temp. Donor 11

Solvent

Yield [%]

[°C]

[equiv.]

1

2

3

4

5

6

7

8

0.3

0.5

–78

–40

–20

–40

1.3

1.6

2.0

CH₂Cl₂

toluene

CH₂Cl₂

toluene

20

30

50

65

60

67

72

82

The selective removal of the levulinoyl ester in 40 gave

alcohol 41 (85%), which was converted to triol 42 by the

acidic hydrolysis of the 4,6-O-isopropylidene acetal (92%).

The Pd/C-mediated benzyl hydrogenolysis and concomitant

allyl reduction of 42 gave the target acetate 5 in 77% yield.

Synthesis of the (E)AB_AcCD and (E)ABCD

Pentasaccharides 5 and 6

Donor 12, easily obtained from the fully protected tetra- Alternatively, the transesterification of 40 in refluxing meth-

saccharide 9 in two steps, served as an exquisite intermedi- anolic MeONa gave diol 43 (94%), which was subsequently

ate in the synthesis of pentasaccharides 5 and 6 (Scheme 7). converted into the corresponding tetraol 44 (66%) upon

–

Scheme 7. Synthesis of pentasaccharides 5 and 6: (a) i. catalytic [Ir(COD){PCH₃(C₆H₅)₂}₂]⁺PF₆, THF, room temp.; ii. I₂, THF/H₂O,

room temp., 90%; (b) CCl₃CN, DBU, DCE, –5 °C, 88%; (c) AllOH, catalytic TMSOTf, CH₂Cl₂, –78 °C, 82%; (d) 1  aqueous NaOH,

MeOH, 71%; (e) TfOH (0.9 equiv.), 4-Å MS, 70 °C, toluene, 78%; (f) hydrazine hydrate, pyridine/AcOH, room temp., 85%; (g) 50%

aqueous TFA, CH₂Cl₂, 0 °C, 92% 42, 70% 44; (h) NaOMe, MeOH, reflux, 94%; (i) Pd/C, H₂, EtOH, 77% 5, 70% 6.

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J. Boutet, L. A. Mulard

FULL PAPER

starting material and the presence of one more polar product. The

reaction mixture was concentrated under reduced pressure.

Chromatography of the residue (Chex/EtOAc, 90:10 Ǟ 75:25) gave

disaccharide 15 (19.7 g, 95%) as a colourless syrup. Compound 15

had R_f= 0.3 (Chex/EtOAc, 7:3). ¹H NMR (400 MHz, CDCl₃): δ

= 7.40–7.10 (m, 25 H, CH_Ph), 5.87 (m, 1 H, CH=), 5.39 (m, 1 H,

H-2_A), 5.28 (m, J_trans= 17.2 Hz, 1 H, =CH₂), 5.19 (d, J_1,2= 5.6 Hz,

1 H, H-1_E), 5.28 (m, J_cis= 10.3 Hz, 1 H, =CH₂), 5.02 (d, J =

11.0 Hz, 1 H, H_Bn), 4.96–4.85 (m, 3 H, H_Bn), 4.78 (d, J_1,2= 1.8 Hz,

1 H, H-1_A), 4.78–4.59 (m, 4 H, H_Bn), 4.50 (d, J = 11.0 Hz, 1 H,

acidic hydrolysis of the 4,6-O-isopropylidene protecting

group. The final debenzylation and allyl reduction of 44

gave the free pentasaccharide 6 (70%).

Conclusions

The synthesis of the two linear tetrasaccharides 1, 2 and

of the four branched pentasaccharides 3–6, representative

of fragments of S. flexneri 3a and X O-Ags, was described.

The overall strategy relied on the use of a common tetrasac-

charide EABC intermediate 9, obtained in 28 synthetic

steps. Indeed, fully protected 9, having an allyl aglycon, a

2_C-acetate, and a 2_A-levulinoyl ester, served as a convenient

precursor, providing easy access to all targets. Thus, starting

from 9, both oligosaccharides 1 and 2 were synthesized in

2 steps. Pentasaccharides 3 and 4 were obtained in 7 steps,

whereas pentasaccharides 5 and 6 were prepared in 13 steps

from the shared intermediate 9. All targets were synthesized

as their propyl glycosides. The overall strategy took advan-

tage of the recently disclosed neutral conditions^[15]allowing

for concomitant benzyl hydrogenolysis, allyl reduction, and

reductive trichloroacetamide conversion to acetamide.

H_Bn), 4.36 (d, J = 12.0 Hz, 1 H, H_Bn), 4.26 (dd, J_2,3= 3.2, J_3,4

=

9.6 Hz, 1 H, H-3_A), 4.18–4.04 (m, 3 H, H_All, H-3_E, H-5_E), 3.97 (m,

1 H, H_All), 3.80 (m, 1 H, H-5_A), 3.77 (pt, J_4,5= 9.2 Hz, 1 H, H-

4_E), 3.64–3.52 (m, 4 H, H-2_E, H-6a_E, H-4_A, H-6b_E), 2.55 (m, 4 H,

2ϫCH_2Lev), 2.09 (s, 3 H, CH_3Lev), 1.40 (d, J_5,6= 6.2 Hz, 3 H, H-

6_A) ppm. ¹³C NMR (100 MHz, CDCl₃): δ = 206.5 (C_Lev), 172.5

(C_Lev), 139.2–138.1 (C_Ph), 134.0 (CH=), 129.0–127.8 (CH_Ph), 117.8

(=CH₂), 97.0 (¹J_C,H= 170.2 Hz, C-1_E), 93.3 (¹J_C,H= 167.5 Hz, C-

1_A), 82.5 (C-3_E), 80.3 (C-4_A), 80.0 (C-2_E), 78.3 (C-4_E), 76.5, 75.9,

75.3, 73.7, 72.8 (5 C, C_Bn), 72.8 (C-3_A), 70.6 (C-5_E), 68.7 (C-6_E),

68.6 (C-2_A), 68.5 (C_All), 68.4 (C-5_A), 38.2 (CH_2Lev), 30.0 (CH_3Lev),

28.4 (CH_2Lev), 18.3 (C-6_A) ppm. HRMS (ESI⁺): calcd. for

C₅₅H₆₂O₁₂[M + Na]⁺937.4139; found 937.4109, calcd. for [M +

NH₄]⁺932.4585; found 932.4542.

(2,3,4,6-Tetra-O-benzyl-α-

D-glucopyranosyl)-(1Ǟ3)-4-O-benzyl-2-

O-levulinoyl-α/β- -rhamnopyranose (16): 1,5-Cyclooctadiene-bis-

L

(methyldiphenylphosphane)iridium hexafluorophosphate (250 mg)

was dissolved in THF (100 mL), and the resulting red solution was

degassed under an argon stream. Hydrogen was bubbled through

the solution, causing the colour to change to yellow. The solution

was then degassed again under an argon stream. A solution of 15

(8.5 g, 9.3 mmol) in THF (15 mL) was added. The mixture was

stirred overnight at room temp. TLC (Chex/EtOAc, 7:3) showed the

complete disappearance of the starting material and the presence of

a single less polar product. The mixture was treated with a solution

of iodine (4.7 g, 18.6 mmol) in THF/H₂O (50 mL, 4:1 v/v) for 1 h

at room temp. TLC (Chex/EtOAc, 7:3 and CH₂Cl₂/MeOH, 98:2)

showed the complete disappearance of the intermediate and the

presence of a single more polar product. Excess iodine was de-

stroyed by the addition of a solution of freshly prepared sodium

bisulfite (5% aqueous, 40 mL). CH₂Cl₂(300 mL) was added, and

the organic phase was washed with brine (3ϫ50 mL), H₂O (3 ϫ

50 mL), dried on Na₂SO₄, filtered, and concentrated to dryness.

Chromatography of the residue (CH₂Cl₂/MeOH, 99:1 Ǟ 9:1) gave

disaccharide 16 (α/β: 7:3, 7.5 g, 92%) as a yellow syrup. Hemiacetal

16 had R_f= 0.2 (CH₂Cl₂/MeOH, 98:2); α anomer: ¹H NMR

(400 MHz, CDCl₃): δ = 7.43–7.14 (m, 25 H, CH_Ph), 5.43 (m, 1 H,

H-2_A), 5.25 (d, J_1,2= 3.4 Hz, 1 H, H-1_E), 5.15 (d, J_1,2= 1.7 Hz, 1

H, H-1_A), 5.07–4.87 (m, 3 H, H_Bn), 4.80 (d, J = 12.2 Hz, 1 H, H_Bn),

4.74–4.59 (m, 3 H, H_Bn), 4.52 (d, J = 11.0 Hz, 1 H, H_Bn), 4.46–

4.36 (d, 2 H, H_Bn), 4.34 (m, 1 H, H-3_A), 4.20–4.10 (m, 2 H, H-3_E,

H-5_E), 4.05 (m, 1 H, H-5_A), 3.78–3.73 (m, 2 H, OH, H-4_E), 3.70–

3.55 (m, 4 H, H-2_E, H-6a_E, H-4_A, H-6b_E), 2.55 (m, 4 H,

2ϫCH_2Lev), 2.11 (s, 3 H, CH_3Lev), 1.41 (d, J_5,6= 6.2 Hz, 3 H, H-

6_A) ppm. ¹³C NMR (100 MHz, CDCl₃), δ = 206.9 (C_Lev), 172.6

Experimental Section

General: TLC was performed with precoated slides of Silica Gel 60

F₂₅₄(Merck). Detection was effected when applicable, with UV

light, and/or by charring in orcinol (35 m) in 4  aqueous sulfuric

acid and ethanol (95:5). Preparative chromatography was per-

formed by elution from columns of Silica Gel 60 (particle size

0.040–0.063 mm). NMR spectra were recorded at 30 °C for solu-

tions in CDCl₃or D₂O (400 MHz for ¹H, 100 MHz for ¹³C). Resid-

ual CHCl₃(δ = 7.28 ppm for H and 77.0 ppm for ¹³C) and HOD

(δ = 4.79 ppm) were used as internal references for solutions in

CDCl₃and D₂O, respectively. Proton signal assignments were made

1

by first-order analysis of the spectra, as well as analysis of 2D H-

¹H correlation maps (COSY). Of the two magnetically non-equiva-

lent geminal protons at C-6, the one resonating at lower field is

denoted H-6a, and the one at higher field is denoted H-6b. The ¹³

C

NMR assignments were supported by 2D ¹³C-¹H correlation maps

(HMBC and HSQC). Interchangeable assignments are marked

with an asterisk in the listing of signal assignments. Sugar residues

are serially lettered according to the lettering of the repeating unit

of the S. flexneri 3a O-Ag and identified by a subscript in the listing

of signal assignments. Electrospray ionisation/time-of-flight (ESI-

TOF) mass spectra were recorded in the positive-ion mode with 1:1

acetonitrile (CH₃CN)/H₂O containing 0.1% formic acid as the ESI-

TOF spectrometer solution. Anhydrous dichloromethane (CH₂Cl₂)

and 1,2-dichloroethane (DCE), delivered on molecular sieves, were

used as such. 4-Å MS were activated before use by heating at

250 °C under vacuum. Additional solvents commonly cited in the

text are abbreviated as Chex (cyclohexane), THF (tetrahydrofuran),

and Tol (toluene).

(C_Lev), 139.2–138.9 (C_Ph), 129.0–127.8 (CH_Ph), 93.0 (¹J_C,H

=

Allyl (2,3,4,6-Tetra-O-benzyl-α-

zyl-2-O-levulinoyl-α- -rhamnopyranoside (15): DMAP (13.9 g,

D

-glucopyranosyl)-(1Ǟ3)-4-O-ben- 169.0 Hz, C-1_E), 92.4 (¹J_C,H= 170.7 Hz, C-1_A), 82.5 (C-3_E), 80.3

L

(C-4_A), 79.9 (C-2_E), 78.3 (C-4_E), 76.5, 75.9, 75.4, 73.6, 73.2 (5 C,

C_Bn), 72.3 (C-3_A), 70.5 (C-5_E), 69.0 (C-2_A), 68.7 (C-6_E), 68.3 (C-

5_A), 38.3 (CH_2Lev), 30.1 (CH_3Lev), 28.6 (CH_2Lev), 18.5 (C-6_A) ppm.

114 mmol, 5 equiv.) was added to a solution of alcohol 14^[26]

(18.6 g, 22.8 mmol) in pyridine (150 mL). The mixture was stirred

at 50 °C, and levulinic anhydride^[30](48.8 g, 228 mmol, 10 equiv.)

in pyridine (150 mL) was added dropwise to the solution. After 3 h,

TLC (Chex/EtOAc, 7:3) showed the complete disappearance of the

1

β anomer: H NMR (400 MHz, CDCl₃): δ = 7.43–7.14 (m, 25 H,

CH_Ph), 5.62 (d, J_2,3= 2.7 Hz, 1 H, H-2_A), 5.38 (d, J_1,2= 3.4 Hz, 1

H, H-1_E), 5.07–4.87 (m, 3 H, H_Bn), 4.80 (d, J = 12.2 Hz, 1 H, H_Bn),

5532

www.eurjoc.org

Eur. J. Org. Chem. 2008, 5526–5542

Shigella flexneri Serotypes 3a and X O-Antigens

4.79 (s, 1 H, H-1_A), 4.74–4.59 (m, 3 H, H_Bn), 4.53 (d, J = 11.0 Hz,

1 H, H_Bn), 4.46–4.36 (d, 2 H, H_Bn), 4.20–4.10 (m, 2 H, H-3_E, H-

5_E), 4.00 (m, 1 H, H-3_A), 3.75 (m, 1 H, H-4_E), 3.70–3.55 (m, 3 H,

H-2_E, H-6a_E, H-6b_E), 3.53 (pt, J_4,5= 9.3 Hz, 1 H, H-4), 3.44 (m,

1 H, H-5_A), 2.55 (m, 4 H, 2ϫCH_2Lev), 2.14 (s, 3 H, CH_3Lev), 1.47

(d, J_5,6= 6.0 Hz, 3 H, H-6_A) ppm. ¹³C NMR (CDCl₃): δ = 208.6

(C_Lev), 173.1 (C_Lev), 139.2–138.9 (C_Ph), 129.0–127.8 (CH_Ph), 93.6

(¹J_C,H= 160.9 Hz, C-1_A), 92.6 (¹J_C,H= 169.1 Hz, C-1_E), 82.4 (C-

3_E), 79.6 (C-4_A), 80.0 (C-2_E), 78.4 (C-4_E), 76.6, 76.0, 75.5 (3 C,

(33% solution in AcOH, 23 mL) was added dropwise to a solution

of the crude peracetate in CH₂Cl₂(100 mL), which was stirred at

room temp. After 5 h, TLC (Chex/EtOAc, 6:4) showed the com-

plete disappearance of the starting material. Volatiles were evapo-

rated under reduced pressure and eliminated by repeated coevapo-

ration with toluene to provide the bromide (70.6 g) as a light yellow

syrup; R_f= 0.35 (Chex/EtOAc, 6:4); α anomer: ¹H NMR

(400 MHz, CDCl₃): δ = 6.26 (d, J_1,2= 0.8 Hz, 1 H, H-1), 5.66 (dd,

J_3,4= 10.2, J_2,3= 3.4 Hz, 1 H, H-3), 5.44 (m, 1 H, H-2), 5.14 (pt,

J_4,5= 10.0 Hz, 1 H, H-4), 4.10 (dq, 1 H, H-5), 2.14, 2.06, 1.99 (3

s, 9 H, H_Ac), 1.25 (d, J_5,6= 6.3 Hz, 3 H, H-6) ppm. ¹³C NMR

(100 MHz, CDCl₃): δ = 170.1, 170.0, 169.9 (3 C, C_Ac), 84.1 (¹J_C,H

= 184.3 Hz, C-1), 72.8 (C-2), 71.5 (C-5), 70.7 (C-4), 68.3 (C-3),

21.1, 21.0, 20.9 (3 C, C_Ac), 17.3 (C-6) ppm. 2,6-Lutidine (28 mL)

was added to a stirred suspension of the crude bromide (70.6 g)

and allyl alcohol (200 mL) in anhydrous CH₂Cl₂(200 mL) contain-

ing 4-Å MS (10 g). The mixture was stirred at room temp. for 12 h,

at which time TLC (Chex/EtOAc, 7:3) indicated the complete dis-

appearance of the stating material. The mixture was diluted with

CH₂Cl₂(100 mL), washed with cold citric acid (3ϫ20 mL), cold

saturated NaHCO₃(2 ϫ 20 mL), and H₂O (50 mL), dried on

Na₂SO₄, filtered, and concentrated under reduced pressure to pro-

vide crude 20 (66.0 g) as a light yellow syrup. Diacetate 20^[37]had

R_f= 0.35 (Chex/EtOAc, 7:3). ¹H NMR (400 MHz, CDCl₃): δ =

C_Bn), 74.2 (C-3_A), 73.6, 73.4 (2 C, C_Bn), 72.3 (C-5_A), 70.5 (C-5_E),

69.0 (C-2_A), 68.8 (C-6_E), 39.1 (CH_2Lev), 30.0 (CH_3Lev), 28.5

(CH_2Lev), 18.5 (C-6_A) ppm. HRMS (ESI⁺): calcd. for C₅₂H₅₈O₁₂

[M + Na]⁺897.3826; found 897.3777, calcd. for [M + NH₄]⁺

892.4272; found 892.4234.

(2,3,4,6-Tetra-O-benzyl-α-

D-glucopyranosyl)-(1Ǟ3)-4-O-benzyl-2-

O-levulinoyl-α/β- -rhamnopyranose Trichloracetimidate (7): Hemi-

L

acetal 16 (7.1 g, 8.1 mmol) was dissolved in DCE (30 mL), placed

under argon, and cooled to –5 °C. Trichloroacetonitrile (4.0 mL,

40.5 mmol, 5 equiv.) and DBU (340 µL, 2.3 mmol, 0.28 equiv.) were

added. The mixture was stirred at –5 °C for 10 min. TLC (Chex/

EtOAc + Et₃N, 7:3) showed the complete disappearance of 16 and

the presence of a single less polar product. The mixture was directly

chromatographed (Chex/EtOAc + 5% Et₃N, 7:3 Ǟ 1:1) to give 7

(8.0 g, 97%) as a yellow syrup. Trichloroacetimidate 7 (α anomer)

1

had R_f= 0.35 (Chex/EtOAc, 7:3). H NMR (400 MHz, CDCl₃): δ 5.85 (m, 1 H, CH=), 5.39 (d, 1 H, J_1,2= 2.4 Hz, H-1), 5.22 (m, 1

= 8.72 (s, 1 H, NH), 7.42–7.14 (m, 25 H, CH_Ph), 6.24 (d, J_1,2

=

H, J_trans= 17.2 Hz, =CH₂), 5.09 (m, 1 H, J_cis= 11.8 Hz, =CH₂),

2.0 Hz, 1 H, H-1_A), 5.61 (m, 1 H, H-2_A), 5.27 (d, J_1,2= 3.4 Hz, 1 5.06 (dd, J_2,3= 3.9 Hz, J_3,4= 9.7 Hz, 1 H, H-3), 5.01 (pt, J_4,5

=

9.9 Hz, 1 H, H-4), 4.56 (dd, J_1,2= 2.4 Hz, J_2,3= 3.9 Hz, 1 H, H-

2), 4.00 (m, 2 H, H_All), 3.49 (dq, 1 H, H-5), 2.07, 2.02 (2 s, 6 H,

H_Ac), 1.72 (s, 3 H, CH_3ortho), 1.20 (d, J_5,6= 6.2 Hz, 3 H, H-6) ppm.

¹³C NMR (100 MHz, CDCl₃): δ = 170.7, 170.3 (2 C, C_Ac), 134.5

(CH=), 124.5 (C_ortho), 116.9 (=CH₂), 97.4 (¹J_C,H= 175.0 Hz, C-

1), 77.0 (C-2), 71.1 (C-3), 70.7 (C-4), 69.5 (C-5), 64.0 (C_All), 24.5

(CH_3ortho), 21.2, 21.0 (2 C, C_Ac), 17.8 (C-6) ppm. Anhydrous

K₂CO₃(1.1 g, 8.0 mmol) was added to a stirred solution of crude

20 (66.0 g) in dry MeOH (250 mL). The mixture was stirred at

room temp. for 6 h, at which time TLC (CH₂Cl₂/MeOH, 9:1) indi-

cated the total conversion of 20 into a single product. Volatiles were

removed under reduced pressure to give diol 21 (49.2 g) as a pale

foam. Diol 21^[37]had R_f= 0.5 (CH₂Cl₂/MeOH, 9:1). ¹H NMR

(400 MHz, CDCl₃): δ = 5.85 (m, 1 H, CH=), 5.39 (d, J_1,2= 2.0 Hz,

H, H-1_E), 5.05–4.99 (m, 2 H, H_Bn), 4.92–4.88 (m, 2 H, H_Bn), 4.80

(d, J = 12.0 Hz, 1 H, H_Bn), 4.74–4.53 (m, 4 H, H_Bn), 4.42 (d, J =

12.2 Hz, 1 H, H_Bn), 4.34 (dd, J_2,3= 3.1, J_3,4= 9.7 Hz, 1 H, H-3_A),

4.16–4.06 (m, 2 H, H-3_E, H-5_E), 4.02 (m, 1 H, H-5_A), 3.79 (pt, J_3,4

= 9.3 Hz, 1 H, H-4_E), 3.71–3.62 (m, 3 H, H-4_A, H-6a_E, H-2_E), 3.52

(m, 1 H, H-6b_E), 2.59 (m, 4 H, 2ϫCH_2Lev), 2.11 (s, 3 H, CH_3Lev),

1.45 (d, J_5,6= 6.2 Hz, 3 H, H-6_A) ppm. ¹³C NMR (100 MHz,

CDCl₃): δ = 206.3 (C_Lev), 172.5 (C_Lev), 160.5 (C=NH), 139.1–137.9

(C_Ph), 129.2–127.8 (CH_Ph), 95.4 (¹J_C,H= 178.9 Hz, C-1_A), 93.4

(¹J_C,H= 168.3 Hz, C-1_E), 91.3 (CCl₃), 82.4 (C-3_E), 79.8 (C-2_E), 79.5

(C-4_A), 78.2 (C-4_E), 76.8, 75.9, 75.4, 73.8, 73.2 (5 C, C_Bn), 72.5 (C-

3_A), 71.4 (C-5_A), 70.8 (C-5_E), 68.4 (C-6_E), 66.9 (C-2_A), 38.2

(CH_2Lev), 30.0 (CH_3Lev), 28.4 (CH_2Lev), 18.4 (C-6_A) ppm.

3,4-Di-O-benzyl-1,2-(allyloxyethylidene)-β-L-rhamnopyranose (22):

1 H, H-1), 5.26 (m, J_trans= 17.2 Hz, 1 H, =CH₂), 5.15 (m, J_cis

=

Acetic anhydride (100 mL) was added dropwise to a solution of -

rhamnose (36.4 g, 200 mmol) in anhydrous pyridine (100 mL). The

resulting mixture was stirred at room temp. under argon. After 1 d,

TLC (CH₂Cl₂/MeOH, 8:2) showed the complete disappearance of

the starting material. The mixture was concentrated under reduced

pressure, and volatiles were eliminated by repeated coevaporation

with toluene to provide the tetra-acetate (66.3 g) as a colourless

syrup. The product had R_f= 0.3 (Chex/EtOAc, 6:4); α anomer: ¹H

NMR (400 MHz, CDCl₃): δ = 5.98 (d, J_1,2= 1.9 Hz, 1 H, H-1),

5.27 (dd, J_3,4= 10.1, J_2,3= 3.5 Hz, 1 H, H-3), 5.22 (m, 1 H, H-2),

5.09 (pt, J_4,5= 9.9 Hz, 1 H, H-4), 3.91 (dq, 1 H, H-5), 2.14, 2.13,

2.05, 1.98 (4 s, 12 H, H_Ac), 1.20 (d, J_5,6= 6.3 Hz, 3 H, H-6) ppm.

¹³C NMR (100 MHz, CDCl₃), δ = 170.5, 170.2, 170.0, 168.6 (4 C,

C_Ac), 91.0 (¹J_C,H= 177.0 Hz, C-1), 71.8 (C-5_β), 70.8 (C-4), 69.1 (C-

3), 69.1 (C-5), 69.0 (C-2), 21.4, 21.2, 21.0, 20.8 (8 C, C_Ac), 17.8 (C-

10.4 Hz, 1 H, =CH₂), 4.48 (m, 1 H, H-2), 4.07 (m, 2 H, H_All), 3.70

(dd, J_3,4= 9.2, J_2,3= 4.0 Hz, 1 H, H-3), 3.42 (pt, J_4,5= 9.2 Hz, 1

H, H-4), 3.30 (dq, 1 H, H-5), 1.71 (s, 3 H, CH_3ortho), 1.31 (d, J_5,6

= 6.0 Hz, 3 H, H-6) ppm. ¹³C NMR (100 MHz, CDCl₃): δ = 134.4

(CH=), 124.0 (C_ortho), 116.8 (=CH₂), 97.7 (¹J_C,H= 175.1 Hz, C-

1), 79.6 (C-2), 72.9 (C-3), 72.7 (C-4), 71.1 (C-5), 64.0 (C_All), 25.5

(CH_3ortho), 17.8 (C-6) ppm. Sodium hydride (60% suspension in oil,

24.0 g, 600 mmol) was added portionwise to a solution of crude 21

(49.2 g) in dry DMF (300 mL), while the temperature was main-

tained below 5 °C. Stirring was continued for 1 h at room temp.,

and benzyl bromide (57.3 mL, 480.0 mmol, 2.4 equiv.) was then

added dropwise to the reaction mixture, which was kept under

strong stirring below 10 °C. The solution was then stirred at room

temp. for 4 h, when TLC (Chex/EtOAc, 8:2) indicated the presence

of a single less polar product. The mixture was cooled to 0 °C, and

MeOH (30 mL) was added dropwise. After 2 h, EtOAc (400 mL)

was added, and the organic phase was washed with H₂O

(3ϫ50 mL), dried on Na₂SO₄, filtered, and concentrated to dry-

ness. Chromatography of the residue (Chex/EtOAc, 9:1 Ǟ 7:3) gave

22 (64.8 g, 76 % from -rhamnose) as a colourless syrup. Com-

pound 22 had R_f= 0.5 (Chex/EtOAc, 7:3). ¹H NMR (400 MHz,

1

6_α), 17.7 (C-6) ppm. β Anomer: H NMR (400 MHz, CDCl₃): δ =

5.84 (d, J_1,2= 1.2 Hz, 1 H, H-1), 5.45 (m, 1 H, H-2), 5.05 (m, 2 H,

H-3, H-4), 3.65 (dq, 1 H, H-5), 2.20, 2.12, 2.07, 1.95 (4 s, 12 H,

H_Ac), 1.26 (d, J_5,6= 6.2 Hz, 3 H, H-6) ppm. ¹³C NMR (100 MHz,

CDCl₃), δ = 170.3, 170.1, 169.9, 168.7 (4 C, C_Ac), 91.0 (¹J_C,H

=

162.6 Hz, C-1), 71.8 (C-5), 71.1, 70.6 (2 C, C-3*, C-4*), 68.9 (C-2),

21.3, 21.1, 20.9, 20.7 (8 C, C_Ac), 17.8 (C-6_α), 17.7 (C-6) ppm. HBr

Eur. J. Org. Chem. 2008, 5526–5542

www.eurjoc.org

5533

J. Boutet, L. A. Mulard

FULL PAPER

CDCl₃): δ = 7.47–7.35 (m, 10 H, CH_Ph), 5.96 (m, 1 H, CH=), 5.33

with H₂O, saturated NaHCO₃(3ϫ20 mL), brine (3ϫ20 mL) and

(m, J_trans= 17.2 Hz, 1 H, =CH₂), 5.31 (d, J_1,2= 1.7 Hz, 1 H, H- H₂O (3ϫ20 mL). The organic layer was dried on Na₂SO₄, filtered,

1), 5.22 (m, J_cis= 10.4 Hz, 1 H, =CH₂), 4.99 (d, J = 10.8 Hz, 1 H, and concentrated under reduced pressure. Chromatography of the

H_Bn), 4.80 (m, 2 H, H_Bn), 4.71 (d, J = 10.8 Hz, 1 H, H_Bn), 4.41 residue (Chex/EtOAc, 85:15 Ǟ 8:2) gave 27 (2.4 g, 89 %) as a

(dd, J_1,2= 1.7, J_2,3= 4.1 Hz, 1 H, H-2), 4.08 (m, 2 H, H_All), 3.73

(dd, J_3,4= 9.1, J_2,3= 4.1 Hz, 1 H, H-3), 3.52 (pt, J_4,5= 9.1 Hz, 1

colourless syrup. Compound 27 had R_f= 0.4 (Chex/EtOAc, 7:3).

¹H NMR (400 MHz, CDCl₃): δ = 7.39–7.30 (m, 10 H, CH_Ph), 5.91

H, H-4), 3.37 (dq, 1 H, H-5), 1.80 (s, 3 H, CH_3ortho), 1.34 (d, J_5,6(m, 1 H, CH=), 5.43 (m, 1 H, H-2), 5.28 (m, J_trans= 15.8 Hz, 1 H,

= 6.2 Hz, 3 H, H-6) ppm. ¹³C NMR (100 MHz, CDCl₃): δ = 138.7– =CH₂), 5.22 (m, J_cis= 10.4 Hz, 1 H, =CH₂), 4.95 (d, J = 10.8 Hz,

138.3 (C_Ph), 135.0 (CH=), 129.0–128.3 (CH_Ph), 124.1 (C_ortho), 116.9 1 H, H_Bn), 4.81 (d, J_1,2= 1.5 Hz, 1 H, H-1), 4.72 (d, J = 11.2 Hz,

(=CH₂), 97.8 (¹J_C,H= 173.6 Hz, C-1), 79.9 (C-4), 79.6 (C-3), 77.5 1 H, H_Bn), 4.65 (d, J = 10.8 Hz, 1 H, H_Bn), 4.55 (d, J = 11.2 Hz,

(C-2), 76.6, 72.6 (2 C, C_Bn), 70.7 (C-5), 64.0 (C_All), 25.4 (CH_3ortho), 1 H, H_Bn), 4.16 (m, 1 H, H_All), 3.99 (m, 2 H, H_All, H-3), 3.81 (m,

18.4 (C-6) ppm. HRMS (ESI⁺): calcd. for C₂₅H₃₀O₆[M + Na]⁺1 H, H-5), 3.46 (pt, J_4,5= 9.4 Hz, 1 H, H-4), 2.74 (m, 4 H,

449.1940; found 449.1940.

2ϫCH_2Lev), 2.19 (s, 3 H, CH_3Lev), 1.37 (d, J_5,6= 6.2 Hz, 3 H, H-

6) ppm. ¹³C NMR (100 MHz, CDCl₃): δ = 206.6 (C_Lev), 172.5

(C_Lev), 138.9–138.5 (C_Ph), 133.9 (CH=), 128.9–128.1 (CH_Ph), 118.0

(=CH₂), 97.1 (¹J_C,H= 169.8 Hz, C-1), 80.5 (C-4), 78.5 (C-3), 75.8,

72.0 (2 C, C_Bn), 69.5 (C-2), 68.4 (C_All), 68.1 (C-5), 38.4 (CH_2Lev),

30.2 (CH_3Lev), 28.6 (CH_2Lev), 18.4 (C-6) ppm. HRMS (ESI⁺):

calcd. for C₂₈H₃₄O₇[M + Na]⁺505.2202; found 505.2201.

Allyl 2-O-Acetyl-3,4-di-O-benzyl-α-L-rhamnopyranoside (23):

TMSOTf (4.5 mL, 25.2 mmol, 0.2 equiv.) was added to a solution

of 22 (53.8 g, 126 mmol) in CH₂Cl₂(300 mL) containing 4-Å MS

(55 g), and the mixture was stirred at 0 °C. The reaction mixture

was stirred for 3 h while the temperature was slowly raised to room

temp. TLC (Chex/EtOAc, 7:3) showed the complete disappearance

of the starting material and the presence of a major less polar prod-

uct. Et₃N (5 mL) was added, and the mixture was filtered. The

filtrate was concentrated to dryness. Chromatography of the resi-

due (Chex/EtOAc, 9:1 Ǟ 7:3) gave 23 (47.5 g, 88%) as a colourless

syrup. Analytical data were identical to those published.^[35]

3,4-Di-O-benzyl-2-O-levulinoyl-α/β-L-rhamnopyranose (28): 1,5-Cy-

clooctadiene-bis(methyldiphenylphosphane)iridium hexafluoro-

phosphate (120 mg) was dissolved in THF (30 mL), and the re-

sulting red solution was degassed under an argon stream. Hydrogen

was bubbled through the solution, causing the colour to change to

yellow. The solution was then degassed again under an argon

stream. A solution of 27 (2.7 g, 5.7 mmol) in THF (8 mL) was

added. The mixture was stirred overnight at room temp. TLC

(Chex/EtOAc, 7:3) showed the complete disappearance of the start-

ing material and the presence of a single less polar product. The

mixture was treated with iodine (2.9 g, 11.3 mmol) in THF/H₂O

(30 mL, 4:1 v/v) for 1 h at room temp. TLC (Chex/EtOAc, 7:3 and

CH₂Cl₂/MeOH, 98:2) showed the complete disappearance of the

intermediate and the presence of a single more polar product. Ex-

cess iodine was destroyed by the addition of a solution of freshly

prepared sodium bisulfite (5% aqueous, 20 mL). CH₂Cl₂(100 mL)

was added, and the organic phase was washed with brine

(3ϫ30 mL) and H₂O (3ϫ30 mL), dried on Na₂SO₄, filtered, and

concentrated to dryness. Chromatography of the residue (CH₂Cl₂/

MeOH, 99:1 Ǟ 9:1) gave compound 28 (α/β: 75:25, 2.4 g, 96%) as

a yellow syrup. Hemiacetal 28 had R_f= 0.25 (CH₂Cl₂/MeOH,

Allyl 3,4-Di-O-benzyl-α-L-rhamnopyranoside (26): Route B

(Scheme 3). Diol 18^[15,39](1.9 g, 6.6 mmol), dibutyltin oxide (1.8 g,

7.2 mmol, 1.1 equiv.), and 3-Å MS in toluene (50 mL) were heated

at reflux for 8 h. The reaction mixture was cooled to room temp.

and transferred to a flask containing caesium fluoride (1.5 mL,

13.1 mmol, 2 equiv.). Volatiles were removed, and DMF (50 mL)

was added to the residue, followed by benzyl bromide (1.6 mL,

13.1 mmol, 2 equiv.). The reaction mixture was then stirred at room

temp. overnight. TLC (Chex/EtOAc, 7:3) showed the complete dis-

appearance of the starting material and the presence of a single

less polar product. The mixture was filtered. CH₂Cl₂(200 mL) was

added, and the organic phase was washed with brine (3ϫ50 mL)

and H₂O (3ϫ50 mL), dried on Na₂SO₄, filtered, and concentrated

to dryness. Chromatography of the residue (Chex/EtOAc, 8:2 Ǟ

1:1) gave 26 (2.2 g, 88%) as a colourless syrup. Analytical data were

identical to those published.^[35,40]

1

98:2); α anomer: H NMR (400 MHz, CDCl₃): δ = 7.37–7.28 (m,

Route C2 (Scheme 3): Crude orthoester 22 (85.2 g), derived from

-rhamnose (36.4 g, 200 mmol), was dissolved in EtOAc (300 mL),

and the organic phase was washed with HCl (10 % aqueous,

3 ϫ 50 mL), brine (3 ϫ 50 mL), and H₂O (3 ϫ 50 mL), dried on

Na₂SO₄, filtered, and concentrated to dryness. The crude material

was added at 0 °C to allyl alcohol (300 mL) containing acetyl chlo-

ride (35.5 mL, 500 mmol, 2.5 equiv.). The reaction mixture was

stirred at 70 °C for 2.5 h and then at 40 °C overnight. The mixture

was neutralized with solid NaHCO₃, filtered through a Celite pad,

and the filtrate was concentrated. Repeated co-evaporation with

toluene gave a brownish syrup, which was purified by column

chromatography (Chex/EtOAc, 8:2 Ǟ 1:1) to give 26 (55.0 g, 72%

from -rhamnose) as a colourless syrup.

10 H, CH_Ph), 5.40 (m, 1 H, H-2), 5.13 (m, 1 H, H-1), 4.95 (d, J =

10.9 Hz, 1 H, H_Bn), 4.75–4.51 (m, 2 H, H_Bn), 4.20 (d, J = 10.7 Hz,

1 H, H_Bn), 4.03–3.98 (m, 2 H, H-3, H-5), 3.43 (pt, J_3,4= 9.4 Hz, 1

H, H-4), 2.73 (m, 4 H, 2ϫCH_2Lev), 2.18 (s, 3 H, CH_3Lev), 1.33 (d,

J_5,6= 6.2 Hz, 3 H, H-6) ppm. ¹³C NMR (100 MHz, CDCl₃), δ =

206.8 (C_Lev), 172.5 (C_Lev), 138.9–138.0 (C_Ph), 128.8–128.0 (CH_Ph),

92.7 (¹J_C,H= 170.3 Hz, C-1), 80.5 (C-4), 77.9 (C-3), 75.7, 72.0 (2

C, C_Bn), 70.0 (C-2), 68.1 (C-5), 38.5 (CH_2Lev), 30.1 (CH_3Lev), 28.6

(CH_2Lev), 18.4 (C-6) ppm; β anomer: ¹H NMR (400 MHz, CDCl₃):

δ = 7.37–7.28 (m, 10 H, CH_Ph), 5.56 (m, 1 H, H-2), 4.95 (d, J =

10.9 Hz, 1 H, H_Bn), 4.77 (d, 1 H, H-1), 4.75–4.51 (m, 2 H, H_Bn),

4.20 (d, J = 10.7 Hz, 1 H, H_Bn), 3.66 (dd, J_2,3= 3.3, J_3,4= 8.9 Hz,

1 H, H-3), 3.38 (m, 1 H, H-5), 3.21 (m, 1 H, H-4), 2.73 (m, 4 H,

2ϫCH_2Lev), 2.20 (s, 3 H, CH_3Lev), 1.38 (d, J_5,6= 5.9 Hz, 3 H, H-

6) ppm. ¹³C NMR (100 MHz, CDCl₃): δ = 208.2 (C_Lev), 173.0

Allyl 3,4-Di-O-benzyl-2-O-levulinoyl-α-L-rhamnopyranoside (27):

DCC (1.8 g, 8.5 mmol, 1.5 equiv.) and levulinic acid (1.1 mL,

10.2 mmol, 1.8 equiv.), dissolved in CH₂Cl₂(120 mL), were added

to a solution of alcohol 26 (2.2 g, 5.7 mmol) and DMAP (3.6 g,

29.3 mmol, 2 equiv.) in CH₂Cl₂(100 mL). The mixture was stirred

overnight at room temp. TLC (CH₂Cl₂/EtOAc, 9:1) showed the

complete disappearance of the starting material and the presence

of one less polar product. Dicyclohexylurea was filtered away, and

the reaction mixture was diluted with CH₂Cl₂and then washed

(C_Lev), 138.9–138.0 (C_Ph), 128.8–128.0 (CH_Ph), 93.5 (¹J_C,H

=

158.8 Hz, C-1β), 80.4 (C-3), 79.9 (C-4) 75.8, 71.9 (2 C, C_Bn), 72.1

(C-5), 70.5 (C-2), 39.0 (CH_2Lev), 30.2 (CH_3Lev), 28.8 (CH_2Lev), 18.4

(C-6) ppm. HRMS (ESI⁺): calcd. for C₂₅H₃₀O₇[M + Na]⁺

465.1889; found 465.1876.

3,4-Di-O-benzyl-2-O-levulinoyl-α-

L-rhamnopyranose Trichloro-

acetimidate (29): Hemiacetal 28 (2.1 g, 4.8 mmol) was dissolved in

5534

www.eurjoc.org

Eur. J. Org. Chem. 2008, 5526–5542

Shigella flexneri Serotypes 3a and X O-Antigens

DCE (15 mL), placed under argon, and cooled to –5 °C. Trichloro-

colourless syrup. Alcohol 8 had R_f= 0.3 (Tol/EtOAc, 8:2). ¹H

acetonitrile (2.4 mL, 24.2 mmol, 5 equiv.) and DBU (202 µL, NMR (400 MHz, CDCl₃): δ = 7.40–7.28 (m, 15 H, CH_Ph), 5.89 (m,

1.3 mmol, 0.28 equiv.) were added. The mixture was stirred at –5 °C

for 10 min. TLC (Chex/EtOAc + Et₃N, 7:3) showed the complete

disappearance of 28 and the presence of a single less polar product.

The mixture was directly chromatographed (Chex/EtOAc + 5%

Et₃N, 7:3 Ǟ 1:1) to give 29 (2.6 g, 92%) as a yellow syrup. Tri-

chloroacetimidate 29 (α anomer) had R_f= 0.4 (Chex/EtOAc, 7:3).

¹H NMR (400 MHz, CDCl₃): δ = 8.68 (s, 1 H, NH), 7.37–7.28 (m,

1 H, CH=), 5.29 (m, J_trans= 17.2 Hz, 1 H, =CH₂), 5.21 (m, J_cis=

10.4 Hz, 1 H, =CH₂), 5.17 (m, 1 H, H-2_C), 5.10 (d, J_1,2= 1.5 Hz,

1 H, H-1_B), 4.88 (d, J = 11.0 Hz, 1 H, H_Bn), 4.80 (d, J_1,2= 1.6 Hz,

1 H, H-1_C), 4.87 (d, J = 11.0 Hz, 1 H, H_Bn), 4.69–4.60 (d, 4 H,

H_Bn), 4.18–4.13 (m, 2 H, H-3_C, H_All), 4.01–3.96 (m, 2 H, H-2_B,

H_All), 3.83–3.74 (m, 3 H, H-3_B, H-5_B, H-5_C), 3.48 (pt, J_3,4= 9.1 Hz,

1 H, H-4_B), 3.46 (pt, J_3,4= 9.4 Hz, 1 H, H-4_C), 2.14 (s, 3 H, H_Ac),

10 H, CH_Ph), 6.20 (d, J_1,2= 1.9 Hz, 1 H, H-1), 5.50 (m, 1 H, H- 1.31 (m, 6 H, H-6_B, H-6_C) ppm. ¹³C NMR (100 MHz, CDCl₃): δ

2), 4.97 (d, J = 10.8 Hz, 1 H, H_Bn), 4.74 (d, J = 11.3 Hz, 1 H, H_Bn),

4.67 (d, J = 10.8 Hz, 1 H, H_Bn), 4.58 (d, J = 11.3 Hz, 1 H, H_Bn),

4.03–3.94 (m, 2 H, H-3, H-5), 3.53 (pt, J_4,5= 9.5 Hz, 1 H, H-4),

= 170.7 (C_Ac), 138.8–138.3 (C_Ph), 133.9 (CH=), 129.4–128.1

(CH_Ph), 117.9 (=CH₂), 101.8 (¹J_C,H= 171.0 Hz, C-1_B), 96.6 (¹J_C,H

= 169.2 Hz, C-1_C), 80.7 (C-4_C), 80.2 (C-4_B), 80.1 (C-3_B), 78.5 (C-

3_C), 75.9, 75.6 (2 C, C_Bn), 72.9 (C-2_C), 72.4 (C_Bn), 69.4 (C-2_B), 68.7

(C-5_B), 68.6 (C_All), 68.1 (C-5_C), 21.5 (C_Ac), 18.3, 18.2 (2 C, C-6_B, C-

2.78 (m, 4 H, 2ϫCH_2Lev), 2.19 (s, 3 H, CH_3Lev), 1.37 (d, J_5,6

=

6.2 Hz, 3 H, H-6) ppm. ¹³C NMR (100 MHz, CDCl₃): δ = 206.4

(C_Lev), 172.2 (C_Lev), 160.7 (C=NH), 138.5–138.0 (C_Ph), 128.8–128.2 6_C) ppm. HRMS (ESI⁺): calcd. for C₃₈H₄₆O₁₀[M + Na]⁺685.2989;

(CH_Ph), 95.5 (¹J_C,H= 178.6 Hz, C-1), 91.3 (CCl₃), 79.7 (C-4), 77.6

found 685.2993, calcd. for [M + NH₄]⁺680.3434; found 680.3472,

(C-3), 76.0, 72.3 (2 C, C_Bn), 71.1 (C-5), 68.2 (C-2), 38.4 (CH_2Lev), calcd. for [M + K]⁺701.2728; found 701.2731.

30.2 (CH_3Lev), 28.5 (CH_2Lev), 18.4 (C-6) ppm.

Allyl (2,3,4,6-Tetra-O-benzyl-α-D-glucopyranosyl)-(1Ǟ3)-(4-O-ben-

Allyl (3,4-Di-O-benzyl-2-O-levulinoyl-α-

L-rhamnopyranosyl)-(1Ǟ3)-

zyl-2-O-levulinoyl-α-L-rhamnopyranosyl)-(1Ǟ2)-(3,4-di-O-benzyl-α-

2-O-acetyl-4-O-benzyl-α- -rhamnopyranoside (31): TMSOTf

L

-rhamnopyranosyl)-(1Ǟ3)-2-O-acetyl-4-O-benzyl-α-L

-rhamno-

(1.1 mL, 6.0 mmol, 0.3 equiv.) was added to a solution of trichloro-

acetimidate 29 (14.2 g, 24.2 mmol, 1.2 equiv.) and acceptor 30

(6.8 g), obtained from 18 (6.0 g, 20.4 mmol),^[38]in toluene (250 mL)

containing 4-Å MS (17 g), and the mixture was stirred at –78 °C.

The reaction mixture was stirred for 1 h, while the temperature was

slowly raised to room temp. TLC (Tol/EtOAc, 8:2) showed the

complete disappearance of the acceptor and the presence of a

major less polar product. Et₃N (2 mL) was added, and the mixture

was filtered and concentrated to dryness. Chromatography of the

residue (Tol/EtOAc, 95:5 Ǟ 85:15) gave 31 (13.3 g, 89 %) as a

colourless syrup. Disaccharide 31 had R_f= 0.5 (Chex/EtOAc, 6:4).

¹H NMR (400 MHz, CDCl₃): δ = 7.39–7.27 (m, 15 H, CH_Ph), 5.89

pyranoside (9): TMSOTf (340 µL, 1.9 mmol, 0.3 equiv.) was added

to a solution of acceptor 8 (4.2 g, 6.3 mmol) and trichloroacetimid-

ate 7 (8.0 g, 7.8 mmol, 1.2 equiv.) in toluene (100 mL) containing

4-Å MS (5.3 g), and the mixture was stirred at –78 °C. The reaction

mixture was stirred for 1 h while the temperature was slowly raised

to room temp. TLC (Tol/EtOAc, 8:2) showed the complete disap-

pearance of the acceptor and the presence of a major less polar

product. Et₃N (1 mL) was added, and the mixture was filtered and

concentrated to dryness. Chromatography of the residue (Tol/

EtOAc, 95:5 Ǟ 85:15) gave 9 (8.8 g, 92%) as a colourless syrup.

Tetrasaccharide 9 had R_f= 0.45 (Chex/EtOAc, 7:3). ¹H NMR

(400 MHz, CDCl₃): δ = 7.42–7.15 (m, 40 H, CH_Ph), 5.89 (m, 1 H,

(m, 1 H, CH=), 5.46 (m, 1 H, H-2_B), 5.29 (m, J_trans= 17.2 Hz, 1 CH=), 5.58 (m, 1 H, H-2_A), 5.31 (m, J_trans= 17.2 Hz, 1 H, =CH₂),

H, =CH₂), 5.21 (m, J_cis= 10.4 Hz, 1 H, =CH₂), 5.17 (m, 1 H, H- 5.28 (d, J_1,2= 3.4 Hz, 1 H, H-1_E), 5.21 (m, J_cis= 10.4 Hz, 1 H,

2_C), 5.05 (d, J_1,2= 1.5 Hz, 1 H, H-1_B), 4.91 (d, J = 11.0 Hz, 1 H, =CH₂), 5.19 (m, 1 H, H-2_C), 5.04 (d, J = 11.0 Hz, 1 H, H_Bn), 5.01

H

_Bn), 4.87 (d, J = 10.9 Hz, 1 H, H_Bn), 4.80 (d, J_1,2= 1.5 Hz, 1 H,

(d, J_1,2= 1.4 Hz, 1 H, H-1_B), 5.00 (d, J_1,2= 1.6 Hz, 1 H, H-1_A),

4.98–4.87 (m, 4 H, H_Bn), 4.84 (d, J_1,2= 1.6 Hz, 1 H, H-1_C), 4.82–

H-1_C), 4.67–4.60 (d, 3 H, H_Bn), 4.45 (d, J = 11.3 Hz, 1 H, H_Bn),

4.20–4.13 (m, 2 H, H-3_C, H_All), 3.98 (m, 1 H, H_All), 3.90 (dd, J_2,34.61 (m, 8 H, H_Bn), 4.57 (d, J = 11.1 Hz, 1 H, H_Bn), 4.52 (d, J =

= 3.3, J_3,4= 9.3 Hz, 1 H, H-3_B), 3.83–3.74 (m, 2 H, H-5_B, H-5_C), 11.0 Hz, 1 H, H_Bn), 4.36 (d, J = 12.0 Hz, 1 H, H_Bn), 4.28 (dd, J_2,3

3.47 (pt, J_3,4= 9.5 Hz, 1 H, H-4_C), 3.43 (pt, J_3,4= 9.4 Hz, 1 H, H- = 3.1, J_3,4= 9.6 Hz, 1 H, H-3_A), 4.19 (m, 1 H, H_All), 4.17–4.07 (m,

4_B), 2.71 (m, 4 H, 2ϫCH_2Lev), 2.18 (s, 3 H, CH_3Lev), 2.14 (s, 3 H, 3 H, H-3_E, H-3_C, H-5_E), 4.04 (m, 1 H, H_All), 3.99 (m, 1 H, H-2_B),

H_Ac), 1.31 (m, 6 H, H-6_C, H-6_B) ppm. ¹³C NMR (100 MHz, 3.90 (m, 1 H, H-5_A), 3.88–3.81 (m, 2 H, H-3_B, H-4_E), 3.79–3.74

CDCl₃): δ = 206.4 (C_Lev), 172.2 (C_Lev), 170.6 (C_Ac), 138.9–138.4 (m, 1 H, H-5_C), 3.73 (m, 1 H, H-5_B), 3.69–3.65 (m, 2 H, H-2_E, H-

(C_Ph), 133.9 (CH=), 128.9–128.0 (CH_Ph), 117.8 (=CH₂), 100.0 6a_E), 3.60–3.56 (m, 2 H, H-6b_E, H-4_A), 3.54 (pt, J_3,4= 9.4 Hz, 1

(¹J_C,H= 169.5 Hz, C-1_B), 96.6 (¹J_C,H= 170.0 Hz, C-1_C), 80.7 (C- H, H-4_B), 3.45 (pt, J_3,4= 9.5 Hz, 1 H, H-4_C), 2.57 (m, 4 H,

4_C), 80.2 (C-4_B), 78.1 (C-3_C), 78.0 (C-3_B), 75.9, 75.6 (2 C, C_Bn),

72.7 (C-2_C), 71.9 (C_Bn), 69.7 (C-2_B), 69.0 (C-5_B), 68.6 (C_All), 68.2

2ϫCH_2Lev), 2.17 (s, 3 H, H_Ac), 2.11 (s, 3 H, CH_3Lev), 1.35 (d, J_5,6

= 6.2 Hz, 3 H, H-6_A), 1.30 (m, 6 H, H-6_C, H-6_B) ppm. ¹³C NMR

(C-5_C), 38.4 (CH_2Lev), 30.2 (CH_3Lev), 28.6 (CH_2Lev), 21.4 (C_Ac), (100 MHz, CDCl₃): δ = 206.6 (C_Lev), 172.1 (C_Lev), 170.8 (C_Ac),

18.3 (2 C, C-6_B, C-6_C) ppm. HRMS (ESI⁺): calcd. for C₄₃H₅₂O₁₂

[M + Na]⁺783.3356; found 783.3364, calcd. for [M + K]⁺778.3802;

found 778.3829.

138.6–138.4 (C_Ph), 134.0 (CH=), 129.0–127.9 (CH_Ph), 117.9

(=CH₂), 101.7 (¹J_C,H= 169.9 Hz, C-1_B), 99.5 (¹J_C,H= 176.6 Hz, C-

1_A), 96.5 (¹J_C,H= 170.6 Hz, C-1_C), 93.3 (¹J_C,H= 168.8 Hz, C-1_E),

82.6 (C-3_E), 80.7 (C-4_B), 80.3 (C-4_C), 80.2 (C-4_A), 79.8 (C-2_E), 79.7

(C-3_B), 79.4 (C-3_C), 78.2 (C-4_E), 76.5, 76.0, 75.9 (3 C, C_Bn), 75.8

(C-2_B), 75.7, 75.4, 73.8, 73.2 (4 C, C_Bn), 72.9 (C-2_C), 72.6 (C-3_A),

72.5 (C_Bn), 70.6 (C-5_E), 69.4 (C-5_B), 69.0 (C-5_A), 68.7 (C_All), 68.6

(C-6_E), 68.4 (C-2_A), 68.1 (C-5_C), 38.3 (CH_2Lev), 30.1 (CH_3Lev), 28.5

(CH_2Lev), 21.5 (C_Ac), 18.4, 18.3, 18.2 (3 C, C-6_A, C-6_B, C-6_C) ppm.

HRMS (ESI⁺): calcd. for C₉₀H₁₀₂O₂₁[M + Na]⁺1541.6812;

found 1541.6769, calcd. for [M + NH₄]⁺1536.7257; found

1536.7207.

Allyl (3,4-Di-O-benzyl-α-

L-rhamnopyranosyl)-(1Ǟ3)-2-O-acetyl-4-

O-benzyl-α- -rhamnopyranoside (8): A solution of hydrazine hy-

L

drate (55 µL, 1.1 mmol, 5 equiv.) in pyridine/AcOH (3:2, v/v, 5 mL)

was added to a solution of fully protected 31 (172 mg, 23 µmol) in

pyridine (1 mL). After 30 min at room temp., TLC (Tol/EtOAc,

8:2) showed the complete disappearance of 31 and the presence of

a major more polar product. H₂O (10 mL) and CH₂Cl₂(50 mL)

were added to the mixture, and the organic phase was washed with

brine (3ϫ10 mL) and H₂O (3ϫ10 mL), dried on Na₂SO₄, filtered,

and concentrated to dryness. Chromatography of the residue (Tol/

EtOAc, 90:10 Ǟ 75:25) gave disaccharide 8 (121 mg, 81%) as a

Allyl (2,3,4,6-Tetra-O-benzyl-α-

D-glucopyranosyl)-(1Ǟ3)-(4-O-ben-

zyl-α- -rhamnopyranosyl)-(1Ǟ2)-(3,4-di-O-benzyl-α-

L

-rhamno-

Eur. J. Org. Chem. 2008, 5526–5542

www.eurjoc.org

5535

J. Boutet, L. A. Mulard

FULL PAPER

pyranosyl)-(1Ǟ3)-2-O-acetyl-4-O-benzyl-α-

L

-rhamnopyranoside

(C-3_C), 75.7 (C-3_A), 73.3 (C-3_E), 72.5 (C-4_C), 72.4 (C-4_E), 72.3 (C-

2_C), 72.1 (C-5_E), 71.8 (C-2_E), 70.7 (C-4_A), 70.3 (C_Pr), 70.2 (C-5_A),

69.8 (2 C, C-4_B, C-5_B), 69.7 (C-3_B), 69.0 (C-5_C), 67.1 (C-2_A), 60.7

(C-6_E), 22.4 (CH₂), 20.7 (C_Ac), 17.2, 17.1, 16.9 (3 C, C-6_A, C-6_B,

C-6_C), 10.3 (CH₃) ppm. HRMS (ESI⁺): calcd. for C₂₉H₅₀O₁₉[M +

Na]⁺725.2844; found 725.2827.

(10): A solution of hydrazine hydrate (88 µL, 1.8 mmol, 5 equiv.)

in pyridine/AcOH (3:2, v/v, 5 mL) was added to a solution of fully

protected 9 (552 mg, 1.8 mmol) in pyridine (1 mL). After 30 min at

room temp., TLC (Tol/EtOAc, 8:2) showed the complete disappear-

ance of the starting material and the presence of a major less polar

product. H₂O (10 mL) and CH₂Cl₂(50 mL) were added to the mix-

ture, and the organic phase was washed with brine (3ϫ10 mL) and

H₂O (3ϫ10 mL), dried on Na₂SO₄, filtered, and concentrated to

dryness. Chromatography of the residue (Tol/EtOAc, 9:1 Ǟ 8:2)

gave disaccharide 10 (456 mg, 89%) as a colourless syrup. Alcohol

Allyl (2,3,4,6-Tetra-O-benzyl-α-

zyl-α- -rhamnopyranosyl)-(1Ǟ2)-(3,4-di-O-benzyl-α-

pyranosyl)-(1Ǟ3)-4-O-benzyl-α- -rhamnopyranoside (32): MeONa

D

-glucopyranosyl)-(1Ǟ3)-(4-O-ben-

L

-rhamno-

L

(0.5  in MeOH, 770 µL, 390 µmol, 1.1 equiv.) was added to a solu-

tion of fully protected 9 (500 mg, 350 µmol) in MeOH (10 mL),

and the mixture was refluxed for 3 h. TLC (Chex/EtOAc, 7:3)

showed the complete disappearance of the starting material and

the presence of a single more polar product. The mixture was neu-

tralized by the addition of Dowex X8-200 ion-exchange resin (H⁺),

and filtered. The evaporation of the filtrate gave a syrup, which was

chromatographed (Chex/EtOAc, 75:25 Ǟ 7:3) to give 32 (461 mg,

95%) as a white foam. Tetrasaccharide 32 had R_f= 0.3 (Chex/

EtOAc, 7:3). ¹H NMR (400 MHz, CDCl₃): δ = 7.42–7.20 (m, 40

H, CH_Ph), 5.93 (m, 1 H, CH=), 5.33 (m, J_trans= 17.2 Hz, 1 H,

=CH₂), 5.23 (m, J_cis= 10.4 Hz, 1 H, =CH₂), 5.18 (s, 1 H, H-1_A),

5.10 (d, J_1,2= 1.5 Hz, 1 H, H-1_B), 5.03–4.98 (m, 3 H, H_Bn), 4.95

(d, J_1,2= 3.3 Hz, 1 H, H-1_E), 4.92–4.87 (m, 3 H, H_Bn), 4.84 (d, J_1,2

= 1.5 Hz, 1 H, H-1_C), 4.79 (d, J = 11.4 Hz, 1 H, H_Bn), 4.75–4.69

(m, 5 H, H_Bn), 4.60–4.56 (m, 2 H, H_Bn), 4.52 (d, J = 11.0 Hz, 1 H,

H_Bn), 4.33 (d, J = 12.0 Hz, 1 H, H_Bn), 4.19 (m, 1 H, H_All), 4.13–

4.07 (m, 5 H, H-2_A, H-3_E, H-2_B, H-3_A, H-2_C), 4.03 (m, 1 H, H_All),

4.01–3.98 (m, 2 H, H-3_C, H-5_E), 3.93–3.87 (m, 3 H, H-3_B, H-5_B,

1

10 had R_f= 0.7 (Tol/EtOAc, 8:2). H NMR (400 MHz, CDCl₃): δ

= 7.43–7.23 (m, 40 H, CH_Ph), 5.89 (m, 1 H, CH=), 5.35 (m, J_trans

= 17.2 Hz, 1 H, =CH₂), 5.25 (m, J_cis= 10.4 Hz, 1 H, =CH₂), 5.22

(m, 1 H, H-2_C), 5.19 (s, 1 H, H-1_A), 5.06 (d, J_1,2= 1.4 Hz, 1 H, H-

1_B), 5.03–4.96 (m, 3 H, H_Bn), 4.97 (d, J_1,2= 3.7 Hz, 1 H, H-1_E),

4.91–4.89 (m, 3 H, H_Bn), 4.81 (d, J_1,2= 1.3 Hz, 1 H, H-1_C), 4.81

(d, J = 10.7 Hz, 1 H, H_Bn), 4.75–4.68 (m, 3 H, H_Bn), 4.66–4.53 (m,

5 H, H_Bn), 4.36 (d, J = 11.9 Hz, 1 H, H_Bn), 4.19 (m, 1 H, H_All),

4.17–4.08 (m, 5 H, H-3_C, H-2_A, H-2_B, H-3_E, H-3_A), 4.03 (m, 1 H,

H_All), 4.01 (m, 1 H, H-5_E), 3.93 (m, 1 H, H-5_A), 3.89 (dd, J_2,3

2.8, J_3,4= 9.4 Hz, 1 H, H-3_B), 3.85 (m, 1 H, H-5_C), 3.79 (pt, J_3,4

=

9.7 Hz, 1 H, H-4_E), 3.74 (m, 1 H, H-5_B), 3.66 (dd, J_1,2= 3.6, J_2,3

= 9.6 Hz, 1 H, H-2_E), 3.57 (pt, J_3,4= 9.3 Hz, 1 H, H-4_A), 3.54–

3.43 (m, 4 H, H-6a_E, H-4_B, H-4_C, H-6b_E), 2.19 (s, 3 H, H_Ac), 1.37

(d, J_5,6= 6.3 Hz, 3 H, H-6_A), 1.35 (d, J_5,6= 6.2 Hz, 3 H, H-6_B),

1.33 (d, J_5,6= 6.2 Hz, 3 H, H-6_C) ppm. ¹³C NMR (100 MHz,

CDCl₃): δ = 170.8 (C_Ac), 138.8–138.6 (C_Ph), 134.0 (CH=), 129.1–

128.1 (CH_Ph), 117.9 (=CH₂), 101.9 (¹J_C,H= 168.4 Hz, C-1_B), 101.3

(¹J_C,H= 173.1 Hz, C-1_A), 96.6 (¹J_C,H= 170.4 Hz, C-1_C), 94.3 (¹J_C,H

= 167.2 Hz, C-1_E), 82.8 (C-3_E), 80.8 (C-4_B), 80.4 (C-4_C), 80.0 (C-

3_B), 79.6 (C-4_A), 79.4 (2 C, C-2_E, C-3_C), 78.2 (C-4_E), 76.8 (C-3_A),

76.0, 75.9, 75.7 (4 C, C_Bn), 75.6 (C-2_B), 75.3, 74.8, 73.8 (3 C, C_Bn),

73.0 (C-2_C), 72.7 (C_Bn), 71.1 (C-5_E), 69.4 (C-5_B), 68.7 (C_All), 68.7

(C-6_E), 68.3 (C-5_A), 68.2 (C-5_C), 67.8 (C-2_A), 21.6 (C_Ac), 18.4, 18.3,

18.2 (3 C, C-6_A, C-6_B, C-6_C) ppm. HRMS (ESI⁺): calcd. for

C₈₅H₉₆O₁₉[M + Na]⁺1443.6444; found 1443.6505, calcd. for [M

+ NH₄]⁺1438.6890; found 1438.6960.

H-5_A), 3.81–3.76 (m, 2 H, H-4_E, H-5_C), 3.65 (dd, J_1,2= 3.5, J_2,3

=

9.7 Hz, 1 H, H-2_E), 3.56 (pt, J_3,4= 9.5 Hz, 1 H, H-4_A), 3.54–3.43

(m, 4 H, H-4_B, H-6a_E, H-4_C, H-6b_E), 2.19 (s, 1 H, OH), 1.37 (d,

J_5,6= 6.2 Hz, 3 H, H-6_A), 1.34 (d, J_5,6= 6.3 Hz, 3 H, H-6_B, H-6_A),

1.30 (d, J_5,6= 6.3 Hz, 3 H, H-6_C) ppm. ¹³C NMR (100 MHz,

CDCl₃): δ = 138.6–138.2 (C_Ph), 134.0 (CH=), 129.0–128.1 (CH_Ph),

117.7 (=CH₂), 101.5 (¹J_C,H= 170.1 Hz, C-1_B), 10143 (¹J_C,H

170.1 Hz, C-1_A), 98.8 (¹J_C,H= 167.2 Hz, C-1_C), 94.3 (¹J_C,H

=

169.2 Hz, C-1_E), 82.8 (C-3_E), 81.5 (C-3_C), 80.7 (C-4_B), 80.2 (C-4_C),

80.0 (C-3_B), 79.6 (C-4_A), 79.3 (C-2_E), 78.2 (C-4_E), 76.8 (C-3_A), 76.0,

75.9, 75.8 (4 C, C_Bn), 75.9 (C-2_B), 75.3, 74.8, 73.8, 72.7 (4 C, C_Bn),

71.3 (C-2_C), 71.1 (C-5_E), 69.6 (C-5_B), 68.4 (C_All), 68.3 (C-6_E), 68.3

(C-5_A), 68.0 (C-5_C), 67.7 (C-2_A), 18.4, 18.3, 18.2 (3 C, C-6_A, C-

6_B, C-6_C) ppm. HRMS (ESI⁺): calcd. for C₈₃H₉₄O₁₈[M + Na]⁺

1401.6338; found 1401.6403, calcd. for [M + NH₄]⁺1396.6783;

found 1396.6854.

Propyl α-

D

-Glucopyranosyl-(1Ǟ3)-α-

L

-rhamnopyranosyl-(1Ǟ2)-α-

-rhamnopyranoside (1):

L

-rhamnopyranosyl-(1Ǟ3)-2-O-acetyl-α-

L

Mono-hydroxy compound 10 (400 mg, 281 µmol) was dissolved in

EtOH (30 mL) and treated with Pd/C (10%, 300 mg), and the sus-

pension was stirred at room temp. overnight under a hydrogen at-

mosphere. TLC (iPrOH/H₂O/NH₃, 4:1:0.5 and Tol/EtOAc, 8:2)

showed that starting material had been transformed into a major

polar product. The suspension was filtered through an Acrodisc

LC (25 mm), and the filtrate was concentrated. Reverse phase

chromatography (H₂O/CH₃CN, 100:0 Ǟ 70:30) of the residue, fol-

lowed by freeze-drying, gave the target tetrasaccharide 1 (150 mg,

Propyl α-

D

-Glucopyranosyl-(1Ǟ3)-α-

L-rhamnopyranosyl-(1Ǟ2)-α-

L

-rhamnopyranosyl-(1Ǟ3)-α-

L

-rhamnopyranoside (2): Diol 32

(360 mg, 261 µmol) was dissolved in EtOH (30 mL), treated with

Pd/C (10%, 300 mg), and the suspension was stirred at room temp.

overnight under a hydrogen atmosphere. TLC (iPrOH/H₂O/NH₃,

4:1:0.5 and Chex/EtOAc, 7:3) showed that the starting material had

been transformed into a major more polar product. The suspension

76%) as a white foam. Tetrasaccharide 1 had R_f= 0.8 (iPrOH/

1

H₂O/NH₃, 4:1:0.5). H NMR (400 MHz, D₂O): δ = 5.09 (d, J_1,2

=

1.0 Hz, 1 H, H-1_B), 5.07 (m, 1 H, H-2_C), 5.00 (d, J_1,2= 3.8 Hz, 1 was filtered through an Acrodisc LC (25 mm), and the filtrate was

H, H-1_E), 4.88 (d, J_1,2= 1.5 Hz, 1 H, H-1_A), 4.72 (d, J_1,2= 1.5 Hz, concentrated. Reverse-phase chromatography (H₂O/CH₃CN, 100:0

1 H, H-1_C), 4.17 (dd, J_2,3= 2.5 Hz, 1 H, H-2_A), 3.91–3.83 (m, 3 H, Ǟ 70:30) of the residue, followed by freeze-drying, gave the target

H-2_B, H-3_C, H-5_E), 3.75–3.61 (m, 7 H, H-3_A, H-5_C, H-6a_E, H-6b_E,

H-3_E, H-5_A, H-3_B), 3.60–3.54 (m, 2 H, H_Pr, H-5_B), 3.49 (pt, J_3,4

tetrasaccharide 2 (140 mg, 81%) as a white foam. Tetrasaccharide

=

2 had R_f= 0.5 (iPrOH/H₂O/NH₃, 4:1:0.5). ¹H NMR (400 MHz,

9.7 Hz, 1 H, H-4_C), 3.48–3.37 (m, 3 H, H-2_E, H-4_A, H_Pr), 3.42 (pt, D₂O): δ = 5.10 (br. s, 1 H, H-1_B), 4.99 (d, J_1,2= 3.8 Hz, 1 H, H-

J_3,4= 10.0 Hz, 1 H, H-4_B), 3.42 (pt, J_3,4= 9.2 Hz, 1 H, H-4_E), 2.07

(s, 3 H, H_Ac), 1.60 (sext, 2 H, CH₂), 1.27 (d, J_5,6= 6.2 Hz, 3 H, H-

6_C), 1.18–1.16 (m, 6 H, H-6_A, H-6_B), 0.88 (t, J = 7.4 Hz, 3 H, CH₃)

1_E), 4.89 (d, J_1,2= 1.5 Hz, 1 H, H-1_A), 4.65 (d, J_1,2= 1.4 Hz, 1 H,

H-1_C), 4.17 (dd, J_2,3= 2.4 Hz, 1 H, H-2_A), 3.95 (m, 1 H, H-2_B)

3.88–3.81 (m, 3 H, H-2_C, H-5_E, H-3_B), 3.73 (dd, J_2,3= 3.0, J_3,4

=

ppm. ¹³C NMR (100 MHz, D₂O): δ = 173.4 (C_Ac), 102.4 (¹J_C,H9.7 Hz, 1 H, H-3_A), 3.71–3.59 (m, 7 H, H-3_C, H-6a_E, H-6b_E, H-5_B,

= 175.5 Hz, C-1_A), 101.2 (¹J_C,H= 173.2 Hz, C-1_B), 97.2 (¹J_C,H

172.3 Hz, C-1_C), 95.8 (¹J_C,H= 167.3 Hz, C-1_E), 78.8 (C-2_B), 76.2 H-4_A, H-4_C, H_Pr, H-4_B), 3.34 (pt, J_3,4= 9.3 Hz, 1 H, H-4_E), 1.50

=

H-3_E, H-5_C, H-5_A), 3.53 (m, 1 H, H_Pr), 3.48–3.37 (m, 5 H, H-2_E,

5536

www.eurjoc.org

Eur. J. Org. Chem. 2008, 5526–5542

Shigella flexneri Serotypes 3a and X O-Antigens

(sext, 2 H, CH₂), 1.18–1.16 (m, 9 H, H-6_A, H-6_B, H-6_C), 0.81 (t, J (1Ǟ3)-4-O-benzyl-α-L-rhamnopyranoside (36): MeONa (0.5  in

= 7.4 Hz, 3 H, CH₃) ppm. ¹³C NMR (100 MHz, D₂O): δ = 102.3 MeOH, 431 µL, 200 µmol, 2 equiv.) was added to a solution of fully

(¹J_C,H= 167.2 Hz, C-1_A), 101.0 (¹J_C,H= 173.1 Hz, C-1_B), 99.8

(¹J_C,H= 169.4 Hz, C-1_C), 95.7 (¹J_C,H= 169.5 Hz, C-1_E), 78.8 (C- ture was stirred for 25 min at room temp. TLC (CH₂Cl₂/MeOH,

2_B), 77.7 (C-3_C), 75.5 (C-3_A), 73.2 (C-3_E), 72.4 (C-4_B), 72.2 (C-4_C), 95:5 and Chex/EtOAc, 6:4) showed the complete disappearance of

72.0 (C-5_E), 71.7 (C-4_A), 70.6 (C-2_E), 70.3 (C-2_C), 70.2 (C-3_B), 69.9 the starting material and the presence of two more polar products.

(C_Pr), 69.7 (C-5_B), 69.6 (C-4_E), 69.5 (C-5_C), 68.9 (C-5_A), 67.0 (C- The mixture was neutralized by the addition of Dowex X8-200 ion-

protected 34 (200 mg, 100 µmol) in MeOH (12 mL), and the mix-

2_A), 60.6 (C-6_E), 22.3 (CH₂), 17.1, 17.0, 16.8 (3 C, C-6_A, C-6_B, C- exchange resin (H⁺), filtered, and the solvents were evaporated to

6_C), 10.2 (CH₃) ppm. HRMS (ESI⁺): calcd. for C₂₇H₄₈O₁₈[M +

Na]⁺683.2739; found 683.2729.

dryness. Chromatography of the residue (Tol/EtOAc, 7:3 Ǟ 1:1)

gave, by order of elution, first triol 35 (103 mg, 56%) and then

tetraol 36 (55 mg, 30%), both as white foams. Triol 35 had R_f=

Allyl (3,4,6-Tri-O-acetyl-2-deoxy-2-trichloroacetamido-β-

pyranosyl)-(1Ǟ2)-[2,3,4,6-tetra-O-benzyl-α- -glucopyranosyl-

(1Ǟ3)]-(4-O-benzyl-α- -rhamnopyranosyl)-(1Ǟ2)-(3,4-di-O-benzyl-

α- -rhamnopyranosyl)-(1Ǟ3)-2-O-acetyl-4-O-benzyl-α- -rhamnopy-

D-gluco-

1

0.3 (CH₂Cl₂/MeOH, 95:5). H NMR (400 MHz, CDCl₃): δ = 7.41

D

(d, J_2,NH= 6.1 Hz, 1 H, NH), 7.39–7.08 (m, 40 H, CH_Ph), 5.94 (m,

1 H, CH=), 5.38 (br. s, 1 H, H-1_A), 5.33 (m, J_trans= 17.2 Hz, 1 H,

=CH₂), 5.23 (m, J_cis= 10.4 Hz, 1 H, =CH₂), 5.19–5.15 (m, 4 H,

H-2_C, H-1_E, 2H_Bn), 5.12–5.01 (m, 2 H, H_Bn), 4.96 (d, J = 10.7 Hz,

1 H, H_Bn), 4.91 (br. s, 1 H, H-1_B), 4.82 (d, J_1,2= 1.5 Hz, 1 H, H-

1_C), 4.80–4.64 (m, 5 H, H_Bn), 4.55 (m, 1 H, H-1_D), 4.54–4.49 (m,

5 H, H_Bn), 4.32 (d, J = 11.9 Hz, 1 H, H_Bn), 4.20 (m, 1 H, H-2_B),

4.17 (m, 1 H, H_All), 4.15 (m, 1 H, H-2_A), 4.13–4.08 (m, 4 H, H-3_E,

H-3_A, H-3_C, H-5_E), 4.11 (m, 1 H, H_All), 3.89–3.82 (m, 3 H, H-4_E,

H-3_B, H-2_E), 3.79–3.69 (m, 4 H, H-2_D, H-5_C, H-5_A, H-5_B), 3.63 (m,

1 H, H-6a_D), 3.52–3.36 (m, 5 H, H-4_B, H-4_A, H-6a_E, H-6b_E, H-

4_C), 3.12 (pt, J_3,4= 9.3 Hz, 1 H, H-4_D), 3.05 (m, 1 H, H-5_D), 2.92

(m, 1 H, H-6b_D), 2.37 (m, 1 H, H-3_D), 2.14 (s, 3 H, H_Ac), 1.34–

1.27 (m, 6 H, H-6_A, H-6_B), 1.28 (d, J_5,6= 6.1 Hz, 3 H, H-6_C) ppm.

¹³C NMR (100 MHz, CDCl₃): δ = 170.8 (C_Ac), 164.9 (C_NTCA),

138.7–138.1 (C_Ph), 134.0 (CH=), 129.5–127.7 (CH_Ph), 118.2

(=CH₂), 102.0 (¹J_C,H= 170.5 Hz, C-1_B), 101.3 (¹J_C,H= 161.0 Hz,

C-1_D), 100.4 (¹J_C,H= 171.2 Hz, C-1_A), 96.5 (¹J_C,H= 171.2 Hz, C-

1_C), 94.6 (¹J_C,H= 169.8 Hz, C-1_E), 92.9 (CCl₃), 83.4 (C-3_E), 80.6

(C-4_B), 80.5 (C-4_E), 80.1 (C-4_C), 80.0 (C-3_C), 79.9 (C-3_B), 79.4 (C-

4_A), 79.0 (C-2_E), 76.6 (C_Bn), 76.5 (C-3_D), 76.1 (C-5_D), 75.9, 75.6,

75.4, 75.3 (5 C, C_Bn), 74.4 (C-3_A), 74.1 (C-2_A), 73.9, 73.2 (2 C,

L

ranoside (34): TMSOTf (28 µL, 160 µmol, 0.5 equiv.) was added to

a solution of acceptor 10 (442 mg, 310 µmol) and trichloroacetim-

idate 11 (370 mg, 620 µmol, 2 equiv.) in toluene (8 mL) containing

4-Å MS (253 mg), and the mixture was stirred at –40 °C. The reac-

tion mixture was stirred for 1 h while the temperature was slowly

raised to room temp. TLC (Tol/EtOAc, 8:2) showed the complete

disappearance of the acceptor and the presence of a major less

polar product. Et₃N (0.1 mL) was added, and the mixture was fil-

tered and concentrated to dryness. Chromatography of the residue

(Tol/EtOAc, 9:1 Ǟ 7:3) gave 34 (469 mg, 82%) as a white foam.

Pentasaccharide 34 had R_f= 0.45 (Tol/EtOAc, 8:2). ¹H NMR

(400 MHz, CDCl₃): δ = 7.43–7.09 (m, 41 H, CH_Ph, NH), 5.89 (m,

1 H, CH=), 5.31 (m, J_trans= 17.2 Hz, 1 H, =CH₂), 5.26 (d, J_1,2

=

3.5 Hz, 1 H, H-1_E), 5.22 (m, J_cis= 10.4 Hz, 1 H, =CH₂), 5.18 (m,

2 H, H_Bn), 5.16 (m, 1 H, H-2_C), 5.19 (d, J_1,2= 1.1 Hz, 1 H, H-1_A),

5.06 (m, 1 H, H-4_D), 5.05 (m, 2 H, H_Bn), 5.00 (d, J_1,2= 1.0 Hz, 1

H, H-1_B), 4.96 (m, 1 H, H_Bn), 4.94 (m, 1 H, H-1_D), 4.88 (pt, J_3,4

=

10.4 Hz, 1 H, H-3_D), 4.81 (d, J_1,2= 1.6 Hz, 1 H, H-1_C), 4.77–4.85

(m, 4 H, H_Bn), 4.62–4.56 (m, 4 H, H_Bn), 4.52–4.47 (m, 2 H, H_Bn),

4.32 (d, J = 11.9 Hz, 1 H, H_Bn), 4.26 (m, 1 H, H-2_D), 4.20 (m, 1

H, H-2_A), 4.18–4.14 (m, 3 H, H_All, H-3_A, H-3_E), 4.12–4.08 (m, 2

H, H-5_E, H-3_C), 4.01 (m, 1 H, H_All), 3.98 (m, 1 H, H-6a_D), 3.96–

3.92 (m, 2 H, H-2_B, H-2_E), 3.87–3.82 (m, 4 H, H-6b_D, H-4_E, H-5A,

C_Bn), 72.9 (C-4_D), 72.8 (C-2_C), 72.1 (C-2_B), 70.4 (C-5_B), 69.7 (C-

5_E), 69.2 (C-5_A), 68.8 (C_All), 68.0 (C-6_E), 67.9 (C-5_C), 62.9 (C-6_D),

58.8 (C-2_D), 21.5 (C_Ac), 18.3, 18.2, 18.1 (3 C, C-6_A, C-6_B, C-6_C)

ppm. HRMS (ESI⁺): calcd. for C₉₃H₁₀₆Cl₃NO₂₄[M + Na]⁺

1748.6068; found 1748.6366, calcd. for [M + NH₄]⁺1743.6514;

found 1743.6545.

H-3_B), 3.79 (m, 1 H, H-5_C), 3.74 (m, 1 H, H-5_B), 3.50 (pt, J_3,4

=

9.3 Hz, 1 H, H-4_B), 3.48 (pt, J_3,4= 9.4 Hz, 1 H, H-4_A), 3.54–3.40

(m, 3 H, H-6a_E, H-6b_E, H-4_C), 2.97 (m, 1 H, H-5_D), 2.15, 2.08,

2.03, 1.90 (4 s, 12 H, H_Ac), 1.34 (d, J_5,6= 6.2 Hz, 3 H, H-6_A), 1.32

(d, J_5,6= 6.2 Hz, 3 H, H-6_B) 1.27 (d, J_5,6= 6.2 Hz, 3 H, H-6_C)

ppm. ¹³C NMR (100 MHz, CDCl₃): δ = 171.0, 170.9, 170.8, 169.6

(4 C, C_Ac), 162.6 (C_NTCA), 139.0–138.0 (C_Ph), 134.0 (CH=), 129.0–

127.8 (CH_Ph), 117.9 (=CH₂), 101.7 (¹J_C,H= 173.2 Hz, C-1_B), 101.3

(¹J_C,H= 171.0 Hz, C-1_A), 101.2 (¹J_C,H= 161.0 Hz, C-1_D), 96.5

(¹J_C,H= 168.8 Hz, C-1_C), 95.1 (¹J_C,H= 164.5 Hz, C-1_E), 93.1

(CCl₃), 82.8 (C-3_E), 80.8 (C-4_B), 80.3 (C-4_C), 80.1 (C-4_A), 79.3 (C-

3_C), 79.2 (C-2_E), 79.1 (C-4_E), 78.9 (C-3_B), 76.4 (C_Bn), 76.0 (C-2_B),

75.9, 75.7, 75.5, 75.3 (4 C, C_Bn), 75.1 (C-3_A), 74.4 (C-2_A), 74.3, 73.8

(2 C, C_Bn), 73.6 (C-3_D), 72.9 (C-2_C), 72.3 (C_Bn), 72.2 (C-5_D), 70.4

(C-5_E), 69.2 (C-5_B), 69.1 (C-5_A), 68.6 (C_All), 68.2 (2 C, C-4_D, C-

6_E), 68.1 (C-5_C), 61.9 (C-6_D), 56.2 (C-2_D), 21.5, 21.0, 20.9, 20.8 (4

C, C_Ac), 18.3, 18.2, 18.1 (3 C, C-6_A, C-6_B, C-6_C) ppm. HRMS

(ESI⁺): calcd. for C₉₉H₁₁₂Cl₃NO₂₇[M + Na]⁺1874.6385; found

1874.6638, calcd. for [M + NH₄]⁺1869.6831; found 1869.6978.

Tetraol 36 had R_f= 0.2 (CH₂Cl₂/MeOH, 95:5). ¹H NMR

(400 MHz, CDCl₃): δ = 7.41 (d, J_2,NH= 6.6 Hz, 1 H, NH), 7.40–

7.11 (m, 40 H, CH_Ph), 5.92 (m, 1 H, CH=), 5.38 (br. s, 1 H, H-1_A),

5.31 (m, J_trans= 17.2 Hz, 1 H, =CH₂), 5.23 (m, J_cis= 10.4 Hz, 1

H, =CH₂), 5.19 (d, J_1,2= 3.5 Hz, 1 H, H-1_E), 5.14–4.98 (m, 4 H,

H_Bn), 4.96 (br. s, 1 H, H-1_B), 4.93 (d, J = 12.3 Hz, 1 H, H_Bn), 4.81

(d, J_1,2= 1.3 Hz, 1 H, H-1_C), 4.80 (d, J = 10.7 Hz, 1 H, H_Bn), 4.74–

4.66 (m, 6 H, H_Bn), 4.55 (m, J_1,2= 4.4 Hz, 1 H, H-1_D), 4.55–4.69

(m, 3 H, H_Bn), 4.37 (d, J = 11.9 Hz, 1 H, H_Bn), 4.20 (m, 1 H, H_All),

4.18–4.10 (m, 5 H, H-2_B, H-2_A, H-3_A, H-3_E, H-5_E), 4.07 (m, 1 H,

H-2_C), 4.11 (m, 1 H, H_All), 3.97 (m, 1 H, H-3_C), 3.92 (m, 1 H, H-

3_B), 3.89–3.83 (m, 3 H, H-4_E, H-2_E, H-2_D), 3.88 (m, 1 H, H-5_B),

3.76 (m, 1 H, H-5_C), 3.72 (m, 1 H, H-5_A), 3.67 (m, 1 H, H-6a_D),

3.56 (pt, J_3,4= 9.4 Hz, 1 H, H-4_B), 3.53–3.39 (m, 4 H, H-6a_E, H-

6b_E, H-4_A, H-4_C), 3.18 (pt, J_3,4= 8.9 Hz, 1 H, H-4_D), 3.13 (m, 1

H, H-5_D), 3.08 (m, 1 H, H-6b_D), 2.48 (m, 1 H, H-3_D), 1.46 (d, J_5,6

Allyl (2-Deoxy-2-trichloroacetamido-β-

[2,3,4,6-tetra-O-benzyl-α- -glucopyranosyl-(1Ǟ3)]-(4-O-benzyl-α-

rhamnopyranosyl)-(1Ǟ2)-(3,4-di-O-benzyl-α- -rhamnopyranosyl)-

(1Ǟ3)-2-O-acetyl-4-O-benzyl-α- -rhamnopyranoside (35) and Allyl

(2-Deoxy-2-trichloroacetamido-β- -glucopyranosyl)-(1Ǟ2)-[2,3,4,6-

tetra-O-benzyl-α- -glucopyranosyl-(1Ǟ3)]-(4-O-benzyl-α-

rhamnopyranosyl)-(1Ǟ2)-(3,4-di-O-benzyl-α- -rhamnopyranosyl)-

D-glucopyranosyl)-(1Ǟ2)- = 6.2 Hz, 3 H, H-6_B), 1.32 (d, J_5,6= 6.2 Hz, 3 H, H-6_A), 1.28 (d,

D

L-

J_5,6= 6.2 Hz, 3 H, H-6_C) ppm. ¹³C NMR (100 MHz, CDCl₃), δ =

164.7 (C_NTCA), 138.7–138.2 (C_Ph), 134.2 (CH=), 129.6–127.7

(CH_Ph), 117.9 (=CH₂), 101.7 (¹J_C,H= 170.5 Hz, C-1_B), 101.4 (¹J_C,H

L

D

= 158.1 Hz, C-1_D), 100.6 (¹J_C,H= 173.4 Hz, C-1_A), 98.8 (¹J_C,H

=

D

L

-

169.8 Hz, C-1_C), 94.7 (¹J_C,H= 168.3 Hz, C-1_E), 93.0 (CCl₃), 83.7

L

(C-3_E), 81.9 (C-3_C), 80.7 (C-4_B), 80.3 (C-3_B), 80.1 (C-4_A), 80.0 (C-

Eur. J. Org. Chem. 2008, 5526–5542

www.eurjoc.org

5537

J. Boutet, L. A. Mulard

FULL PAPER

4_C), 79.4 (C-4_E), 79.1 (C-2_E), 76.6 (C_Bn), 76.5 (C-3_D), 76.2 (C-5_D),

75.8, 75.7, 75.4, 75.3 (5 C, C_Bn), 74.4 (C-3_A), 74.3 (C-2_A), 73.9, 73.4

H-5_B, H-3_C), 3.73–3.65 (m, 5 H, H-2_D, H-6b_D, H-5_A, H-2_E, H-5_C),

3.61 (m, 1 H, H_Pr), 3.51 (pt, J_3,4= 9.6 Hz, 1 H, H-4_C), 3.47 (m, 1

(2 C, C_Bn), 72.8 (C-4_D), 72.7 (C-2_B), 71.3 (C-2_C), 70.4 (C-5_E), 69.7 H, H_Pr), 3.45 (m, 1 H, H-4_E), 3.43 (m, 1 H, H-4_B), 3.41–3.3 (m, 3

(C-5_B), 69.3 (C-5_A), 68.4 (C-6_E), 68.2 (C_All), 67.9 (C-5_C), 63.0 (C- H, H-5_D, H-4_D, H-3_D), 3.32 (pt, J_3,4= 9.7 Hz, 1 H, H-4_A), 2.06 (s,

6_D), 58.8 (C-2_D), 18.4, 18.3, 18.2 (3 C, C-6_A, C-6_B, C-6_C) ppm.

3 H, H_NAc), 1.57 (sext, 2 H, CH₂), 1.25 (m, 6 H, H-6_B, H-6_C), 1.22

HRMS (ESI⁺): calcd. for C₉₁H₁₀₄Cl₃NO₂₃[M + Na]⁺1706.5962; (d, J_5,6= 6.2 Hz, 3 H, H-6_A), 0.88 (t, J = 7.4 Hz, 3 H, CH₃) ppm.

found 1706.6206, calcd. for [M + NH₄]⁺1701.6409; found

1701.6571.

¹³C NMR (100 MHz, D₂O): δ = 174.8 (C_NAc), 102.3 (¹J_C,H

163.2 Hz, C-1_D), 101.7 (¹J_C,H= 170.5 Hz, C-1_A), 101.0 (¹J_C,H

172.7 Hz, C-1_B), 99.9 (¹J_C,H= 169.8 Hz, C-1_C), 95.0 (¹J_C,H

=

Propyl (2-Acetamido-2-deoxy-β-

glucopyranosyl-(1Ǟ3)]-α- -rhamnopyranosyl-(1Ǟ2)-α-

pyranosyl-(1Ǟ3)-2-O-acetyl-α- -rhamnopyranoside (3): Triol 35

D

-glucopyranosyl)-(1Ǟ2)-[α-

D-

170.5 Hz, C-1_E), 78.9 (C-2_B), 77.8 (C-3_C), 76.3 (C-4_D), 74.6 (C-3_D),

74.5 (C-2_A), 74.2 (C-3_A), 73.5 (C-3_E), 72.6 (C-4_B), 72.5 (C-4_C), 71.8

(C-5_E), 71.7 (C-2_E), 71.2 (C-4_A), 70.4 (C-2_C), 70.2 (2 C, C-5_D, C-

3_B), 70.0 (C_Pr), 69.9 (2 C, C-5_A, C-4_E), 69.5 (C-5_B), 69.0 (C-5_C),

61.1 (C-6_D), 60.7 (C-6_E), 56.0 (C-2_D), 23.0 (C_NAc), 22.4 (CH₂), 17.2,

17.1, 16.9 (3 C, C-6_A, C-6_B, C-6_C), 10.3 (CH₃) ppm. HRMS (ESI⁺):

calcd. for C₃₅H₆₁NO₂₃[M + Na]⁺886.3532; found 886.3522, calcd.

for [M + H]⁺864.3713; found 864.3726.

L

-rhamno-

L

(310 mg, 180 µmol) was dissolved in EtOH (20 mL), treated with

Pd/C (10%, 300 mg), and the suspension was stirred at room temp.

for 2 d under a hydrogen atmosphere (45 bar). TLC (iPrOH/H₂O/

NH₃, 4:1:0.5 and CH₂Cl₂/MeOH, 95:5) showed that the starting

material had been transformed into a more polar product. The sus-

pension was filtered through an Acrodisc LC (25 mm), and the fil-

trate was concentrated. Reverse phase chromatography (H₂O/

CH₃CN, 100:0 Ǟ 70:30) of the residue, followed by freeze-drying,

gave the target pentasaccharide 3 (119 mg, 74%) as a white foam.

Pentasaccharide 3 had R_f= 0.2 (iPrOH/H₂O/NH₃, 4:1:0.5). ¹H

NMR (400 MHz, D₂O): δ = 5.09 (br. s, 1 H, H-1_B), 5.08 (m, 1 H,

H-2_C), 5.07 (d, J_1,2= 3.5 Hz, 1 H, H-1_E), 4.99 (d, J_1,2= 1.5 Hz, 1

H, H-1_A), 4.73 (br. s, 1 H, H-1_C), 4.72 (d, J_1,2= 8.5 Hz, 1 H, H-

1_D), 4.33 (m, 1 H, H-2_A), 3.95 (m, 1 H, H-5_E), 3.93 (m, 1 H, H-

2_B), 3.88 (dd, J_2,3= 3.4, J_3,4= 9.6 Hz, 1 H, H-3_C), 3.83–3.79 (m, 2

H, H-3_A, H-6a_D), 3.76–3.68 (m, 4 H, H-3_E, H-5_C, H-6a_E, H-6b_E),

3.66–3.52 (m, 7 H, H-6b_D, H-2_D, H-3_B, H-5_A, H-2_E, H_Pr, H-5_B),

3.49 (pt, J_3,4= 9.7 Hz, 1 H, H-4_C), 3.42 (m, 1 H, H_Pr), 3.38 (m, 1

H, H-4_E), 3.37–3.29 (m, 4 H, H-4_B, H-5_D, H-4_D, H-3_D), 3.26 (pt,

J_3,4= 9.7 Hz, 1 H, H-4_A), 2.08 (s, 3 H, H_Ac), 2.00 (s, 3 H, H_NAc),

1.53 (sext, 2 H, CH₂), 1.22 (d, J_5,6= 6.2 Hz, 3 H, H-6_C), 1.17 (m,

J_5,6= 6.4 Hz, 3 H, H-6_B), 1.15 (d, J_5,6= 6.2 Hz, 3 H, H-6_A), 0.83

(t, J = 7.4 Hz, 3 H, CH₃) ppm. ¹³C NMR (100 MHz, D₂O): δ =

174.8 (C_NAc), 173.3 (C_Ac), 102.2 (¹J_C,H= 163.9 Hz, C-1_D), 101.7

(¹J_C,H= 173.4 Hz, C-1_A), 101.0 (¹J_C,H= 172.7 Hz, C-1_B), 97.1

(¹J_C,H= 172.0 Hz, C-1_C), 94.9 (¹J_C,H= 170.5 Hz, C-1_E), 78.7 (C-

2_B), 76.3 (C-4_D), 76.0 (C-3_C), 74.5 (C-3_D), 74.4 (C-2_A), 74.1 (C-3_A),

73.4 (C-3_E), 72.5 (C-4_C), 72.3 (C-2_C), 72.2 (C-4_B), 71.7 (C-5_E), 71.6

(C-2_E), 71.1 (C-4_A), 70.2 (C_Pr), 70.1 (C-5_D), 69.8 (C-3_B), 69.7 (3 C,

C-4_E, C-5_A, C-5_B), 68.9 (C-5_C), 61.0 (C-6_D), 60.6 (C-6_E), 56.0 (C-

2_D), 23.0 (C_NAc), 22.3 (CH₂), 20.7 (C_Ac), 17.2, 17.0, 16.8 (3 C, C-

6_A, C-6_B, C-6_C), 10.2 (CH₃) ppm. HRMS (ESI⁺): calcd. for

C₃₇H₆₃NO₂₄[M + Na]⁺928.3638; found 928.3652, calcd. for [M +

H]⁺906.3818; found 906.3831.

(2,3,4,6-Tetra-O-benzyl-α-

O-levulinoyl-α- -rhamnopyranosyl)-(1Ǟ2)-(3,4-di-O-benzyl-α-

rhamnopyranosyl)-(1Ǟ3)-2-O-acetyl-4-O-benzyl-α/β- -rhamno-

D-glucopyranosyl)-(1Ǟ3)-(4-O-benzyl-2-

L

L-

L

pyranose (37): 1,5-Cyclooctadiene-bis(methyldiphenylphosphane)-

iridium hexafluorophosphate (25 mg) was dissolved in THF

(30 mL), and the resulting red solution was degassed under an ar-

gon stream. Hydrogen was bubbled through the solution, causing

the colour to change to yellow. The solution was then degassed

again under an argon stream. A solution of allyl glycoside 9 (1.9 g,

1.2 mmol) in THF (15 mL) was added. The mixture was stirred

overnight at room temp. TLC (Tol/EtOAc, 9:1) showed the com-

plete disappearance of the starting material and the presence of a

single less polar product. The mixture was treated with iodine

(340 mg, 2.5 mmol) in THF/H₂O (10 mL, 4:1 v/v) for 1 h at room

temp. TLC (Tol/EtOAc, 9:1 and CH₂Cl₂/MeOH, 99:1) showed the

complete disappearance of the intermediate and the presence of a

single more polar product. Excess iodine was destroyed by the ad-

dition of a solution of freshly prepared sodium bisulfite (5% aque-

ous, 5 mL). CH₂Cl₂(50 mL) was added, and the organic phase was

washed with brine (3 ϫ 20 mL) and H₂O (3 ϫ 20 mL), dried on

Na₂SO₄, filtered, and concentrated to dryness. Chromatography of

the residue (CH₂Cl₂/MeOH, 100:0 Ǟ 99:1) gave tetrasaccharide 37

(1.7 g, 90 %) as a yellow syrup. Hemiacetal 37 had R_f= 0.25

(CH₂Cl₂/MeOH, 99:1). ¹H NMR (400 MHz, CDCl₃): δ = 7.42–

7.15 (m, 40 H, CH_Ph), 5.56 (m, 1 H, H-2_A), 5.26 (d, J_1,2= 3.4 Hz,

1 H, H-1_E), 5.19 (m, 2 H, H-2_C, H-1_C), 5.04 (d, J = 10.9 Hz, 1 H,

H_Bn), 5.01 (d, J_1,2= 1.2 Hz, 1 H, H-1_B), 4.97 (d, J_1,2= 1.4 Hz, 1

H, H-1_A), 4.96–4.81 (m, 14 H, H_Bn), 4.35 (d, J = 12.0 Hz, 1 H,

H_Bn), 4.28 (dd, J_2,3= 3.0, J_3,4= 9.6 Hz, 1 H, H-3_A), 4.17–3.97 (m,

Propyl (2-Acetamido-2-deoxy-β-

glucopyranosyl-(1Ǟ3)]-α- -rhamnopyranosyl-(1Ǟ2)-α-

pyranosyl-(1Ǟ3)-α- -rhamnopyranoside (4): Tetraol 36 (240 mg,

D

-glucopyranosyl)-(1Ǟ2)-[α-

D-

L

-rhamno 5 H, H-3_C, H-3_E, H-5_E, H-5_C, H-2_B), 3.88 (m, 1 H, H-5_A), 3.86–

L

3.78 (m, 2 H, H-3_B, H-4_E), 3.71 (m, 1 H, H-5_B), 3.67–3.63 (m, 2

H, H-2_E, H-6a_E), 3.56 (pt, J_3,4= 9.3 Hz, 1 H, H-4_A), 3.55 (m, 1 H,

H-6b_E), 3.51 (pt, J_3,4= 9.4 Hz, 1 H, H-4_B), 3.43 (pt, J_3,4= 9.4 Hz,

1 H, H-4_C), 3.13 (m, 1 H, OH), 2.53 (m, 4 H, 2ϫCH_2Lev), 2.16 (s,

3 H, H_Ac), 2.10 (s, 3 H, CH_3Lev), 1.34 (d, J_5,6= 6.2 Hz, 3 H, H-

6_A), 1.29 (d, J_5,6= 6.1 Hz, 3 H, H-6_B), 1.27 (d, J_5,6= 6.2 Hz, 3 H,

H-6_C) ppm. ¹³C NMR (100 MHz, CDCl₃): δ = 206.7 (C_Lev), 172.1

(C_Lev), 170.9 (C_Ac), 139.1–138.8 (C_Ph), 129.1–127.9 (CH_Ph), 101.6

(¹J_C,H= 171.8 Hz, C-1_B), 99.5 (¹J_C,H= 169.9 Hz, C-1_A), 93.2 (¹J_C,H

143 µmol) was dissolved in EtOH (15 mL), treated with Pd/C

(10%, 250 mg), and the suspension was stirred at room temp. for

1 d under a hydrogen atmosphere (45 bar). TLC (iPrOH/H₂O/NH₃,

4:1:0.5 and CH₂Cl₂/MeOH, 95:5) showed that the starting material

had been transformed into a more polar product. The suspension

was filtered through an Acrodisc LC (25 mm), and the filtrate was

concentrated. Reverse phase chromatography (H₂O/CH₃CN, 100:0

Ǟ 70:30) of the residue, followed by freeze-drying, gave the target

4 (91 mg, 74%) as a white foam. Pentasaccharide 4 had R_f= 0.15 = 169.6 Hz, C-1_E), 92.0 (¹J_C,H= 170.2 Hz, C-1_C), 82.5 (C-3_E), 80.6

(iPrOH/H₂O/NH₃, 4:1:0.5). ¹H NMR (400 MHz, D₂O): δ = 5.17

(br. s, 1 H, H-1_B), 5.14 (d, J_1,2= 3.6 Hz, 1 H, H-1_E), 5.06 (d, J_1,2

= 1.2 Hz, 1 H, H-1_A), 4.78 (d, J_1,2= 8.5 Hz, 1 H, H-1_D), 4.73 (br.

(C-4_B), 80.2 (C-4_A), 80.1 (C-4_C), 79.7 (C-2_E), 79.6 (C-3_B), 78.8 (C-

3_C), 78.1 (C-4_E), 76.6, 76.0 (2 C, C_Bn), 75.8 (C-2_B), 75.8, 75.7, 75.4,

74.0 (4 C, C_Bn), 73.2 (C-2_C), 73.1 (C_Bn), 72.6 (C-3_A), 72.4 (C_Bn),

s, 1 H, H-1_C), 4.39 (m, 1 H, H-2_A), 4.03 (m, 1 H, H-2_B), 4.00 (m, 70.5 (C-5_E), 69.3 (C-5_B), 69.0 (C-5_A), 68.6 (C-6_E), 68.4 (C-2_A), 68.1

1 H, H-5_E), 3.95 (m, 1 H, H-2_C), 3.90 (m, 1 H, H-3_B), 3.89–3.85 (C-5_C), 38.3 (CH_2Lev), 30.1 (CH_3Lev), 28.5 (CH_2Lev), 21.5 (C_Ac),

(m, 2 H, H-3_A, H-6a_D), 3.82–3.72 (m, 5 H, H-3_E, H-6a_E, H-6b_E, 18.4, 18.3, 18.2 (3 C, C-6_A, C-6_B, C-6_C) ppm. HRMS (ESI⁺): calcd.

5538

www.eurjoc.org

Eur. J. Org. Chem. 2008, 5526–5542

Shigella flexneri Serotypes 3a and X O-Antigens

for C₈₇H₉₈O₂₁[M + Na]⁺1501.6498; found 1501.6665, calcd. for

[M + NH₄]⁺1496.6945; found 1496.7114.

(ESI⁺): calcd. for C₁₆H₂₂NO₇Cl₃[M + Na]⁺468.0359; found

468.0359.

(2,3,4,6-Tetra-O-benzyl-α-

O-levulinoyl-α- -rhamnopyranosyl)-(1Ǟ2)-(3,4-di-O-benzyl-α-

rhamnopyranosyl)-(1Ǟ3)-2-O-acetyl-4-O-benzyl-α/β- -rhamno 39 (1.0 g, 2.2 mmol) in methanol (40 mL), and the suspension was

D

-glucopyranosyl)-(1Ǟ3)-(4-O-benzyl-2-

Allyl 2-Acetamido-2-deoxy-4,6-O-isopropylidene-β-

D-glucopyrano-

L

L-

side (13):^[25]NaOH (1 , 6.7 mL, 6.7 mmol, 3 equiv.) was added to

L

pyranose Trichloroacetimidate (12): Hemiacetal 37 (1.5 g, 1.0 mmol)

was dissolved in DCE (10 mL), placed under argon, and cooled to

–5 °C. Trichloroacetonitrile (520 µL, 5.2 mmol, 5 equiv.) and DBU

(42 µL, 280 µmol, 0.28 equiv.) were added. The mixture was stirred

at –5 °C for 10 min. TLC (Chex/EtOAc + Et₃N, 7:3) showed the

complete disappearance of 37 and the presence of a single less polar

product. The mixture was directly chromatographed (Chex/EtOAc

+ 5% Et₃N, 8:2 Ǟ 7:3) to give donor 12 (1.5 g, 88%) as a yellow

stirred at room temp. After 1 d, TLC (CH₂Cl₂/MeOH, 9:1) showed

the presence of a single more polar product. Acetic anhydride

(1 mL) was added dropwise to the suspension, and after 3 h, the

volatiles were evaporated. Chromatography of the residue (CH₂Cl₂/

MeOH, 98:2 Ǟ 9:1) gave 13 (476 mg, 71%) as a white foam. Ac-

ceptor 13 had R_f= 0.5 (CH₂Cl₂/MeOH, 9:1). ¹H NMR (400 MHz,

CDCl₃): δ = 5.90 (m, 1 H, CH=), 5.74 (d, J_NH,2= 4.6 Hz, 1 H,

NH_NAc), 5.32 (m, J_trans= 17.2 Hz, 1 H, =CH₂), 5.26 (m, J_cis

=

syrup. Trichloroacetimidate 12 (α anomer) had R_f= 0.35 (Chex/

EtOAc, 7:3). H NMR (400 MHz, CDCl₃): δ = 8.73 (s, 1 H, NH), H, H_All), 4.10 (m, 1 H, H_All), 4.00–3.93 (m, 2 H, H-3, H-6A), 3.82

10.4 Hz, 1 H, =CH₂), 4.68 (d, J_1,2= 8.3 Hz, 1 H, H-1), 4.37 (m, 1

1

7.37–7.14 (m, 40 H, CH_Ph), 6.24 (d, J_1,2= 1.9 Hz, 1 H, H-1_C), 5.56

(dd, J_1,2= 2.7, J_1,2= 4.9 Hz, 1 H, H-2_A), 5.32 (dd, J_1,2= 2.2, J_1,2

= 3.1 Hz, 1 H, H-2_C), 5.26 (d, J_1,2= 3.4 Hz, 1 H, H-1_E), 5.04 (d,

J = 10.9 Hz, 1 H, H_Bn), 5.03 (d, J_1,2= 1.3 Hz, 1 H, H-1_B), 4.99 (d,

J_1,2= 1.6 Hz, 1 H, H-1_A), 4.97–4.79 (m, 6 H, H_Bn), 4.73–4.49 (m,

8 H, H_Bn), 4.35 (d, J = 12.0 Hz, 1 H, H_Bn), 4.25 (dd, J_2,3= 3.1,

J_3,4= 9.6 Hz, 1 H, H-3_A), 4.19 (dd, J_2,3= 3.3, J_3,4= 9.5 Hz, 1 H,

H-3_C), 4.14 (pt, J_3,4= 9.3 Hz, 1 H, H-3_E), 4.08 (m, 1 H, H-5_E),

3.98–3.88 (m, 3 H, H-2_B, H-5_C, H-5_A), 3.86–3.81 (m, 2 H, H-3_B,

H-4_E), 3.73 (m, 1 H, H-5_B), 3.67–3.64 (m, 2 H, H-2_E, H-6a_E), 3.58

(m, 1 H, H-6b_E), 3.57 (pt, J_3,4= 9.6 Hz, 1 H, H-4_A), 3.54 (pt, J_3,4

= 9.5 Hz, 1 H, H-4_C), 3.52 (pt, J_3,4= 9.4 Hz, 1 H, H-4_B), 2.55 (m,

4 H, 2ϫCH_2Lev), 2.19 (s, 3 H, H_Ac), 2.10 (s, 3 H, CH_3Lev), 1.34 (d,

J_5,6= 6.3 Hz, 3 H, H-6_A), 1.31 (d, J_5,6= 6.1 Hz, 3 H, H-6_B), 1.28

(d, J_5,6= 6.0 Hz, 3 H, H-6_C) ppm. ¹³C NMR (100 MHz, CDCl₃):

δ = 206.7 (C_Lev), 172.2 (C_Lev), 170.3 (C_Ac), 160.5 (C=NH), 139.1–

138.8 (C_Ph), 129.1–127.9 (CH_Ph), 101.5 (¹J_C,H= 178.7 Hz, C-1_B),

99.8 (¹J_C,H= 174.2 Hz, C-1_A), 94.6 (¹J_C,H= 178.5 Hz, C-1_C), 93.3

(¹J_C,H= 170.0 Hz, C-1_E), 93.3 (CCl₃), 82.5 (C-3_E), 80.6 (C-4_C), 80.1

(C-4_A), 79.8 (C-4_B), 79.6 (C-2_E), 79.4 (C-3_B), 78.2 (C-3_C), 78.1 (C-

4_E), 76.7 (C_Bn), 76.4 (C-2_B), 76.0, 75.9, 75.8, 75.4, 73.8, 73.2 (6 C,

(m, 1 H, H-6B), 3.61 (pt, J_3,4= 9.4 Hz, 1 H, H-4), 3.49 (m, 1 H,

H-2), 3.31 (ddd, J_5,6b= 5.4, J_4,5= 10.0 Hz, 1 H, H-5), 2.07 (s, 3 H,

H_NAc), 1.55 (s, 3 H, Hi_Pr), 1.47 (s, 3 H, Hi_Pr) ppm. ¹³C NMR

(100 MHz, CDCl₃): δ = 162.7 (C_NAc), 134.2 (CH=), 117.8 (=CH₂),

100.9 (¹J_C,H= 162.1 Hz, C-1), 100.1 (Ci_Pr), 74.6 (C-4), 71.8 (C-3),

70.5 (C_All), 67.5 (C-5), 62.4 (C-6), 57.9 (C-2), 29.4 (Ci_Pr), 23.7

(C_NAc), 19.5 (Ci_Pr) ppm. HRMS (ESI⁺): calcd. for C₁₄H₂₃NO₆[M

+ Na]⁺324.123; found 321.1412, calcd. for [M + H]⁺302.1604;

found 302.1599.

Allyl (2,3,4,6-Tetra-O-benzyl-α-

zyl-2-O-levulinoyl-α- -rhamnopyranosyl)-(1Ǟ2)-(3,4-di-O-benzyl-α-

-rhamnopyranosyl)-(1Ǟ3)-(2-O-acetyl-4-O-benzyl-α- -rhamno-

pyranosyl)-(1Ǟ3)-2-acetamido-2-deoxy-4,6-O-isopropylidene-β-

D-glucopyranosyl)-(1Ǟ3)-(4-O-ben-

L

D

-

glucopyranoside (40): TfOH (45 µL, 515 µmol, 0.9 equiv.) was

added to a solution of acceptor 13 (432 mg, 1.4 mmol, 2.5 equiv.)

and trichloroacetimidate 12 (926 mg, 571 µmol) in toluene (25 mL)

containing 4-Å MS (253 mg), and the mixture was stirred at 0 °C.

The reaction mixture was stirred for 1 h at 75 °C. TLC (Tol/EtOAc,

7:3) showed the presence of a major new product. Et₃N (0.2 mL)

was added, and the mixture was filtered. The filtrate was concen-

trated to dryness. Chromatography of the residue (Tol/EtOAc, 7:3

Ǟ 6:4) gave 40 (780 mg, 78%) as a white foam. Pentasaccharide 40

C_Bn), 72.6 (C-3_A), 72.6 (C_Bn), 71.3 (C-2_C), 71.0 (C-5_C), 70.6 (C-5_E),

69.5 (C-5_B), 68.9 (C-5_A), 68.5 (C-6_E), 68.4 (C-2_A), 38.3 (CH_2Lev),

30.1 (CH_3Lev), 28.5 (CH_2Lev), 21.4 (C_Ac), 18.4, 18.3, 18.2 (3 C, C-

6_A, C-6_B, C-6_C) ppm.

1

had R_f= 0.45 (Tol/EtOAc, 6:4). H NMR (400 MHz, CDCl₃): δ =

7.42–7.10 (m, 40 H, CH_Ph), 5.89 (m, 2 H, NH, CH=), 5.56 (dd,

J_1,2= 2.2 Hz, 1 H, H-2_A), 5.30 (m, J_trans= 17.2 Hz, 1 H, =CH₂),

5.25 (d, J_1,2= 3.4 Hz, 1 H, H-1_E), 5.21 (m, J_cis= 10.4 Hz, 1 H,

=CH₂), 5.15 (d, J_1,2= 8.3 Hz, 1 H, H-1_D), 5.07 (dd, J_1,2= 1.6, J_1,2

= 3.2 Hz, 1 H, H-2_C), 5.02 (d, J = 11.0 Hz, 1 H, H_Bn), 4.98 (d, J_1,2

= 1.5 Hz, 1 H, H-1_A), 4.96 (d, J_1,2= 1.3 Hz, 1 H, H-1_B), 4.97–4.86

(m, 4 H, H_Bn), 4.80 (br. s, 1 H, H-1_C), 4.78 (d, J = 11.9 Hz, 1 H,

H_Bn), 4.73 (d, J = 11.5 Hz, 1 H, H_Bn), 4.67 (d, J = 13.4 Hz, 1 H,

H_Bn), 4.62 (d, J = 12.3 Hz, 1 H, H_Bn), 4.61–4.53 (m, 6 H, H_Bn),

4.50 (d, J = 11.0 Hz, 1 H, H_Bn), 4.39 (pt, J_3,4= 9.2 Hz, 1 H, H-

3_D), 4.31 (m, 1 H, H_All), 4.26 (dd, J_2,3= 3.0, J_3,4= 9.6 Hz, 1 H,

H-3_A), 4.15–4.06 (m, 3 H, H-3_E, H_All, H-5_E), 4.04–3.94 (m, 4 H,

H-5_C, H-3_C, H-2_B, H-6a_D), 3.88 (dq, J_4,5= 9.5 Hz, 1 H, H-5_A),

Allyl 3-O-Acetyl-2-deoxy-4,6-O-isopropylidene-2-trichloroacet-

amido-β-D-glucopyranoside (39): TMSOTf (360 µL, 2.0 mmol,

0.2 equiv.) was added to a solution of 38^[24](5.5 g, 10.1 mmol) and

allyl alcohol (1.4 mL, 20.1 mmol, 2 equiv.) in CH₂Cl₂(70 mL) con-

taining 4-Å MS (3.9 g), and the mixture was stirred at –78 °C. The

reaction mixture was stirred for 30 min. TLC (Tol/EtOAc, 8:2)

showed the complete disappearance of the starting material and

the presence of a major less polar product. Et₃N (1 mL) was added.

The mixture was filtered, and the filtrate was concentrated to dry-

ness. Chromatography of the residue (Tol/EtOAc, 9:1 Ǟ 75:25) gave

39 (3.7 g, 82%) as a white foam. Allyl glycoside 39 had R_f= 0.45

1

(Tol/EtOAc, 1:1). H NMR (400 MHz, CDCl₃): δ = 7.65 (d, J_NH,23.84–3.77 (m, 3 H, H-3_B, H-4_E, H-6b_D), 3.71–3.62 (m, 3 H, H-5_B,

= 9.6 Hz, 1 H, NH_NTCA), 5.81 (m, 1 H, CH=), 5.38 (pt, J_2,3

=

H-6a_E, H-2_E), 3.60–3.52 (m, 3 H, H-6b_E, H-4_A, H-4_D), 3.50 (pt,

J_3,4= 9.4 Hz, 1 H, H-4_B), 3.40 (m, 1 H, H-5_D), 3.38 (pt, J_3,4

9.6 Hz, 1 H, H-4_C), 3.49 (m, J_2,NH= 8.0 Hz, 1 H, H-2_D), 2.53 (m,

4 H, 2ϫCH_2Lev), 2.11 (s, 3 H, H_NAc), 2.09 (s, 3 H, CH_3Lev), 2.04

9.3 Hz, 1 H, H-3), 5.25 (m, J_trans= 17.2 Hz, 1 H, =CH₂), 5.15 (m,

J_cis= 9.2 Hz, 1 H, =CH₂), 4.62 (d, J_1,2= 8.3 Hz, 1 H, H-1), 4.29

(m, 1 H, H_All), 4.18 (m, 1 H, H-2), 4.10 (m, 1 H, H_All), 3.99 (dd,

J_5,6a= 5.5, J_6a,6b= 10.8 Hz, 1 H, H-6A), 4.17 (m, 1 H, H-6B), 3.81

=

(s, 3 H, H_Ac), 1.50 (s, 3 H, Hi_Pr), 1.42 (s, 3 H, Hi_Pr), 1.35 (d, J_5,6

=

(pt, J_3,4= 9.5 Hz, 1 H, H-4), 3.75 (ddd, J_4,5= 9.9 Hz, 1 H, H-5), 6.2 Hz, 3 H, H-6_A), 1.26 (d, J_5,6= 6.2 Hz, 3 H, H-6_B), 1.20 (d, J_5,6

2.08 (s, 3 H, H_Ac), 1.51 (s, 3 H, Hi_Pr), 1.38 (s, 3 H, Hi_Pr) ppm. ¹³C = 6.2 Hz, 3 H, H-6_C) ppm. ¹³C NMR (100 MHz, CDCl₃), δ = 206.1

NMR (100 MHz, CDCl₃): δ = 172.0 (C_Ac), 162.7 (C_NTCA), 133.7

(CH=), 118.0 (=CH₂), 100.9 (¹J_C,H= 158.1 Hz, C-1), 100.1 (Ci_Pr), 133.8 (CH=), 129.1–127.4 (CH_Ph), 117.5 (=CH₂), 101.2 (¹J_C,H

93.1 (CCl₃), 72.5 (C-3), 72.1 (C-4), 70.6 (C_All), 67.4 (C-5), 62.2 (C- 169.4 Hz, C-1_B), 99.4 (Ci_Pr), 99.0 (¹J_C,H= 173.7 Hz, C-1_A), 98.9

6), 56.1 (C-2), 29.1 (Ci_Pr), 21.1 (C_Ac), 19.3 (Ci_Pr) ppm. HRMS

(¹J_C,H= 162.7 Hz, C-1_D), 97.6 (¹J_C,H= 171.0 Hz, C-1_C), 92.8 (¹J_C,H

(C_Lev), 171.7 (C_Lev), 171.3 (C_Ac), 170.5 (C_NAc), 138.8–137.6 (C_Ph),

=

Eur. J. Org. Chem. 2008, 5526–5542

www.eurjoc.org

5539

J. Boutet, L. A. Mulard

= 167.6 Hz, C-1_E), 82.1 (C-3_E), 80.2 (C-4_B), 79.8 (C-4_A), 79.6 (C- aqueous, 4 mL) was added, at 0 °C, to a solution of pentasaccha-

FULL PAPER

4_C), 79.5 (C-2_E), 79.4 (2 C, C-3_B, C-3_C), 77.8 (C-4_E), 76.3 (C_Bn),

76.1 (C-3_D), 75.6, 75.5, 75.3 (3 C, C_Bn), 75.2 (C-2_B), 75.1, 75.0, 73.4

ride 41 (400 mg, 240 µmol) in CH₂Cl₂(10 mL), and the biphasic

mixture was stirred vigorously at room temp. for 1 h. TLC (Tol/

(3 C, C_Bn), 73.1 (C-4_D), 72.8 (C-2_C), 72.8 (C_Bn), 72.2 (C-3_A), 72.0 EtOAc, 4:6) showed the complete disappearance of the starting ma-

(C_Bn), 70.5 (C_All), 70.3 (C-5_E), 69.0 (C-5_B), 68.6 (C-5_A), 68.3 (C- terial and the presence of a major more polar product. Repeated

6_E), 68.0 (C-2_A), 67.6 (C-5_C), 67.0 (C-5_D), 62.3 (C-6_D), 59.6 (C-2_D), coevaporation with toluene and chromatography of the residue

37.9 (CH_2Lev), 29.7 (CH_3Lev), 29.1 (Ci_Pr), 28.1 (CH_2Lev), 23.5 (C_Ac), (CH₂Cl₂/MeOH, 98:2 Ǟ 9:1) provided triol 42 (360 mg, 92%) as a

21.2 (C_NAc), 19.3 (Ci_Pr), 18.0, 17.9, 17.8 (3 C, C-6_A, C-6_B, C-6_C) white foam. The triol had R_f= 0.55 (CH₂Cl₂/MeOH, 95:5). ¹H

ppm. HRMS (ESI⁺): calcd. for C₁₀₁H₁₁₉NO₂₆[M + Na]⁺

1784.7917; found 1784.7917, calcd. for [M + H]⁺1762.8098; found

1762.8038, calcd. for [M + NH₄]⁺1779.8364; found 1779.8289.

NMR (400 MHz, CDCl₃): δ = 7.42–7.13 (m, 40 H, CH_Ph), 6.09 (m,

1 H, NH), 5.91 (m, 1 H, CH=), 5.27 (m, 1 H, =CH₂), 5.22 (br. s,

1 H, H-1_A), 5.18 (m, J_cis= 10.4 Hz, 1 H, =CH₂), 5.16 (m, 2 H, H-

1_C, H-2_C), 5.11 (d, J_1,2= 8.3 Hz, 1 H, H-1_D), 5.10–4.88 (m, 8 H,

H-1B, H-1_E, H_Bn), 4.83–4.49 (m, 9 H, H_Bn), 4.35–4.29 (m, 2 H,

Allyl (2,3,4,6-Tetra-O-benzyl-α-

benzyl-α- -rhamnopyranosyl)-(1Ǟ2)-(3,4-di-O-benzyl-α-

pyranosyl)-(1Ǟ3)-(2-O-acetyl-4-O-benzyl-α- -rhamnopyranosyl)-

(1Ǟ3)-2-acetamido-2-deoxy-4,6-O-isopropylidene-β- -gluco-

D

-glucopyranosyl)-(1Ǟ3)-(4-O-

L

-rhamno-

H_All, H_Bn), 4.14–3.96 (m, 9 H, H-3_E, H_All, H-2_A, H-5_C, H-3_A, H-

L

3_C, H-5_E, H-5_A), 3.87–3.73 (m, 4 H, H-3_B, H-6a_D, H-6b_D, H-4_E),

3.68 (m, 1 H, H-5_B), 3.65 (dd, J_2,3= 3.6, J_3,4= 9.6 Hz, 1 H, H-2_E),

3.59 (d, J_4,5= 9.3 Hz, 1 H, H-4_A), 3.54–3.36 (m, 6 H, H-6a_E, H-

5_D, H-4_B, H-4_C, H-6b_E, H-4_D), 3.16 (m, 1 H, H-2_D), 2.14 (s, 3 H,

H_Ac), 2.10 (s, 3 H, H_NAc), 1.25 (m, 9 H, H-6_A, H-6_B, H-6_C) ppm.

¹³C NMR (100 MHz, CDCl₃): δ = 170.5 (2 C, C_NAc, C_Ac), 138.6–

137.5 (C_Ph), 133.9 (CH=), 128.6–127.5 (CH_Ph), 117.7 (=CH₂),

101.3 (C-1_B), 100.8 (¹J_C,H= 168.2 Hz, 2 C, C-1_A, C-1_C), 98.4 (¹J_C,H

= 169.0 Hz, C-1_D), 94.0 (¹J_C,H= 166.3 Hz, C-1_E), 82.4 (C-3_E), 80.2

(C-4_C), 79.4–79.1 (4 C, C-4_B, C-3_B, C-3_C, C-4_A), 79.0 (C-2_E), 77.8

(C-4_E), 76.5 (C-3_A), 75.9, 75.6, 75.4 (3 C, C_Bn), 75.1 (C-4_D), 74.9,

74.5, 74.5, 73.5, 73.5 (5 C, C_Bn), 72.2 (C-2_C), 70.8 (C-5_E), 70.5

(C_All), 70.3 (C-5_D), 69.0 (C-5_B), 68.0 (C-5_A), 67.9 (C-6_E), 67.4 (2 C,

C-5_C, C-2_A), 62.3 (C-6_D), 59.7 (C-2_D), 23.2 (C_NAc), 21.1 (C_Ac), 18.3,

17.9, 17.7 (3 C, C-6_A, C-6_B, C-6_C) ppm. HRMS (ESI⁺): calcd. for

C₉₃H₁₀₉NO₂₄[M + Na]⁺1646.7238; found 1646.7377, calcd. for

[M + H]⁺1624.7418; found 1624.7565, calcd. for [M + NH₄]⁺

1641.7683; found 1641.7848.

D

pyranoside (41): Hydrazine hydrate (200 µL, 4.0 mmol, 10 equiv.) in

pyridine/AcOH (3:2, v/v, 20 mL) was added to a fully protected 40

(720 mg, 4.0 mmol) in pyridine (3 mL). After 30 min at 0 °C, TLC

(Tol/EtOAc, 6:4) showed the complete disappearance of the start-

ing material and the presence of a major less polar product. H₂O

(20 mL) and CH₂Cl₂(100 mL) were added to the mixture, and the

organic phase was washed with brine (3 ϫ 30 mL) and H₂O

(3ϫ30 mL), dried on Na₂SO₄, filtered, and concentrated to dry-

ness. Chromatography of the residue (Tol/EtOAc, 9:1 Ǟ 75:25) gave

alcohol 41 (580 mg, 85%) as a white foam. Pentasaccharide 41 had

R_f= 0.45 (Tol/EtOAc, 6:4). ¹H NMR (400 MHz, CDCl₃): δ = 7.44–

7.18 (m, 40 H, CH_Ph), 5.96 (d, J_NH,2= 6.8 Hz, 1 H, NH), 5.91 (m,

1 H, CH=), 5.32 (m, J_trans= 17.2 Hz, 1 H, =CH₂), 5.24 (m, J_cis

=

10.4 Hz, 1 H, =CH₂), 5.20 (br. s, 1 H, H-1_A), 5.16 (d, J_1,2= 8.4 Hz,

1 H, H-1_D), 5.12 (dd, J_1,2= 1.6, J_1,2= 3.2 Hz, 1 H, H-2_C), 5.03 (d,

J_1,2= 1.3 Hz, 1 H, H-1_B), 5.02–4.95 (d, J = 11.0 Hz, 3 H, H_Bn),

4.95 (d, J_1,2= 3.9 Hz, 1 H, H-1_E), 4.91–4.86 (m, 3 H, H_Bn), 4.86

(br. s, 1 H, H-1_C), 4.84–4.81 (m, 2 H, H_Bn), 4.72–4.70 (m, 3 H,

H_Bn), 4.63–4.58 (m, 4 H, H_Bn), 4.53 (d, J = 11.0 Hz, 1 H, H_Bn),

4.41 (pt, J_3,4= 9.3 Hz, 1 H, H-3_D), 4.34 (m, 1 H, H_All), 4.15–3.96

(m, 9 H, H_All, H-2_B, H-3_E, H-2_A, H-3_A, H-5_C, H-3_C, H-5_E, H-6a_D),

Propyl α-

-rhamnopyranosyl-(1Ǟ3)-(2-o-acetyl-α-

(1Ǟ3)-2-acetamido-2-deoxy-β- -glucopyranoside (5): Pentasaccha-

D

-Glucopyranosyl-(1Ǟ3)-α-

L

-rhamnopyranosyl-(1Ǟ2)-α-

L

-rhamnopyranosyl)-

D

ride 42 (230 mg, 142 µmol) was dissolved in EtOH (15 mL), treated

with Pd/C (10%, 200 mg), and the suspension was stirred at room

temp. for 2 d under a hydrogen atmosphere. TLC (iPrOH/H₂O/

NH₃, 4:1:0.5 and CH₂Cl₂/MeOH, 95:5) showed that 42 had been

transformed into a more polar product. The suspension was filtered

through an Acrodisc LC (25 mm), and the filtrate was concen-

trated. Reverse phase chromatography (H₂O/CH₃CN, 100:0 Ǟ

70:30) of the residue, followed by freeze-drying, gave the target

pentasaccharide 5 (100 mg, 77%) as a white foam. Pentasaccharide

3.91 (dq, J_4,5= 9.6 Hz, 1 H, H-5_A), 3.88 (dd, J_2,3= 2.7, J_3,4

9.4 Hz, 1 H, H-3_B), 3.83 (m, 1 H, H-6b_D), 3.80 (pt, J_3,4= 10.1 Hz,

1 H, H-4_E), 3.72 (dq, J_4,5= 9.4 Hz, 1 H, H-5_B), 3.66 (dd, J_2,3

=

3.6, J_3,4= 9.6 Hz, 1 H, H-2_E), 3.62–3.56 (m, 2 H, H-4_D, H-4_A),

3.54–3.40 (m, 5 H, H-6a_E, H-4_B, H-6b_E, H-4_C, H-5_D) 3.16 (m, J_2,NH

= 8.0 Hz, 1 H, H-2_D), 2.16 (s, 3 H, H_Ac), 2.09 (s, 3 H, H_NAc), 1.54

(s, 3 H, Hi_Pr), 1.46 (s, 3 H, Hi_Pr), 1.38 (d, J_5,6= 6.2 Hz, 3 H, H-

6_A), 1.33 (d, J_5,6= 6.5 Hz, 3 H, H-6_B), 1.26 (d, J_5,6= 6.2 Hz, 3 H,

H-6_C) ppm. ¹³C NMR (100 MHz, CDCl₃): δ = 171.3 (C_NAc), 170.5

(C_Ac), 138.7–137.6 (C_Ph), 133.9 (CH=), 129.1–127.7 (CH_Ph), 117.6

(=CH₂), 101.6 (¹J_C,H= 171.0 Hz, C-1_B), 100.8 (¹J_C,H= 175.0 Hz,

1

5 had R_f= 0.35 (iPrOH/H₂O/NH₃, 4:1:0.5). H NMR (400 MHz,

D₂O): δ = 5.03 (br. s, 1 H, H-1_B), 5.00 (d, J_1,2= 3.7 Hz, 1 H, H-

1_E), 4.90 (m, 2 H, H-1_A, H-2_C), 4.76 (br. s, 1 H, H-1_C), 4.40 (d, J_1,2

= 8.6 Hz, 1 H, H-1_D), 4.17 (m, 1 H, H-2_A), 3.99 (m, J_5,6= 6.3, J_4,5

= 9.8 Hz, 1 H, H-5_C), 3.92 (dd, J_2,3= 5.2 Hz, 1 H, H-2_B), 3.88–

3.80 (m, 3 H, H-5E, H-3_C, H-6a_D), 3.76–3.61 (m, 9 H, H-3_A, H_Pr,

H-2_D, H-6a_E, H-6b_E, H-3_E, H-5_A, H-3_B, H-6b_D), 3.51–3.40 (m, 7

H, H-4_C, H-4_A, H-2_E, H-4_D, H_Pr, H-3_D, H-5_B), 3.38–3.31 (m, 3 H,

H-4_E, H-4_B, H-5_D), 2.07 (s, 3 H, H_Ac), 1.94 (s, 3 H, H_NAc), 1.49

(sext, J = 7.1 Hz, 2 H, CH₂), 1.19–1.14 (m, 9 H, H-6_A, H-6_B, H-

6_C), 0.76 (t, J = 7.4 Hz, 3 H, CH₃) ppm. ¹³C NMR (100 MHz,

D₂O): δ = 174.4 (C_NAc), 173.0 (C_Ac), 101.9 (¹J_C,H= 175.6 Hz, C-

1_A), 101.0 (¹J_C,H= 172.4 Hz, C-1_B), 100.6 (¹J_C,H= 162.7 Hz, C-

1_D), 98.5 (¹J_C,H= 173.4 Hz, C-1_C), 95.3 (¹J_C,H= 170.3 Hz, C-1_E),

82.4 (C-3_D), 78.0 (C-2_B), 76.2 (C-3_C), 76.0 (C-5_D), 75.2 (C-3_A), 72.9

C-1_A), 99.4 (Ci_Pr), 99.0 (¹J_C,H= 160.7 Hz, C-1_D), 97.6 (¹J_C,H

=

170.8 Hz, C-1_C), 94.0 (¹J_C,H= 166.2 Hz, C-1_E), 82.4 (C-3_E), 80.4

(C-4_B), 79.7 (2 C, C-4_C, C-3_B), 79.5 (C-3_C), 79.3 (C-4_A), 79.0 (C-

2_E), 77.8 (C-4_E), 76.5 (C-3_A), 76.2 (C-3_D), 75.7, 75.6, 75.4, 75.3,

75.0 (5 C, C_Bn), 74.9 (C-2_B), 74.5, 73.5 (2 C, C_Bn), 73.1 (C-4_D), 72.9

(C-2_C), 72.3 (C_Bn), 70.8 (C-5_E), 70.5 (C_All), 69.0 (C-5_B), 68.0 (C-

6_E), 67.9 (C-5_A), 67.6 (C-5_C), 67.4 (C-2_A), 67.1 (C-5_D), 62.3 (C-6_D),

59.7 (C-2_D), 29.2 (Ci_Pr), 23.6 (C_NAc), 21.2 (C_Ac), 19.3 (Ci_Pr), 18.0,

17.9, 17.8 (3 C, C-6_A, C-6_B, C-6_C) ppm. HRMS (ESI⁺): calcd. for

C₉₆H₁₁₃NO₂₄[M + Na]⁺1686.7550; found 1686.7488, calcd. for

[M + H]⁺1664.7731; found 1664.7709, calcd. for [M + NH₄]⁺

1681.7997; found 1681.7993.

Allyl (2,3,4,6-Tetra-O-benzyl-α-

zyl-α- -rhamnopyranosyl)-(1Ǟ2)-(3,4-di-O-benzyl-α-

pyranosyl)-(1Ǟ3)-(2-O-acetyl-4-O-benzyl-α- -rhamnopyranosyl)-

(1Ǟ3)-2-acetamido-2-deoxy-β-

D-glucopyranosyl)-(1Ǟ3)-(4-O-ben- (C-3_E), 72.2 (C_Pr), 72.2 (C-2_C), 71.8 (C-4_B), 71.6 (C-5_E), 71.6 (C-

L

-rhamno-

4_C), 71.4 (C-4_A), 70.2 (C-4_D), 69.9 (C-3_B), 69.4 (C-2_E), 69.3 (C-4_E),

69.3 (C-5_A), 68.8 (C-5_C), 68.3 (C-5_B), 66.6 (C-2_A), 60.7 (C-6_D), 60.2

L

D

-glucopyranoside (42): TFA (50% (C-6_E), 55.2 (C-2_D), 22.2 (C_NAc), 22.1 (CH₂), 20.2 (C_Ac), 16.7 (C-

5540

www.eurjoc.org

Eur. J. Org. Chem. 2008, 5526–5542

Shigella flexneri Serotypes 3a and X O-Antigens

6_A), 16.5 (C-6_B), 16.3 (C-6_C), 9.6 (CH₃) ppm. HRMS (ESI⁺): calcd.

for C₃₇H₆₃NO₂₄[M + Na]⁺928.3638; found 928.3635, calcd. for

[M + H]⁺906.3818; found 906.3821, calcd. for [M + K]⁺944.3377;

found 944.3279.

(m, 4 H, H-3_E, H-2_B, H-2_A, H-3_A), 4.00–3.91 (m, 5 H, H-2_C, H-

5_E, H-5_B, H-3_B, H-6a_D), 3.88–3.82 (m, 4 H, H-3_C, H-5_C, H-5_A, H-

6b_D), 3.77 (pt, J_3,4= 9.7 Hz, 1 H, H-4_E), 3.63 (dd, J_2,3= 9.6 Hz, 1

H, H-2_E), 3.56–3.49 (m, 3 H, H-4_A, H-6a_E, H-4_B), 3.47–3.37 (m, 4

H, H-4_D, H-5_D, H-4_C, H-6b_E), 3.15 (m, 1 H, H-2_D), 1.97 (s, 3 H,

H_NAc), 1.34–1.28 (m, 9 H, H-6_A, H-6_B, H-6_C) ppm. ¹³C NMR

(100 MHz, CDCl₃), δ = 170.6 (C_NAc), 138.6–137.5 (C_Ph), 133.9

Allyl (2,3,4,6-Tetra-O-benzyl-α-

zyl-α- -rhamnopyranosyl)-(1Ǟ2)-(3,4-di-O-benzyl-α-

pyranosyl)-(1Ǟ3)-(4-O-benzyl-α- -rhamnopyranosyl)-(1Ǟ3)-2-acet-

amido-2-deoxy-4,6-O-isopropylidene-β- -glucopyranoside (43):

D

-glucopyranosyl)-(1Ǟ3)-(4-O-ben-

L

-rhamno-

L

(CH=), 128.6–127.5 (CH_Ph), 117.5 (=CH₂), 101.5 (¹J_C,H

171.3 Hz, C-1_C), 100.9 (¹J_C,H= 168.8 Hz, C-1_A), 100.9 (¹J_C,H

168.8 Hz, C-1_B), 98.7 (¹J_C,H= 163.1 Hz, C-1_D), 94.0 (¹J_C,H

=

D

MeONa (0.5  in MeOH, 830 µL, 414 µmol, 1 equiv.) was added

to a solution of compound 40 (729 mg, 414 µmol) in MeOH

(20 mL), and the mixture was refluxed for 3 h. TLC (Tol/EtOAc,

1:1) showed the complete disappearance of the starting material

and the presence of a single more polar product. The mixture was

neutralized by addition of Dowex X8-200 ion-exchange resin (H⁺)

and filtered. Evaporation of the filtrate gave a syrup, which was

chromatographed (Tol/EtOAc, 1:1 Ǟ 4:6) to give 43 (631 mg, 94%)

as a white foam. Pentasaccharide 43 had R_f= 0.2 (Tol/EtOAc, 1:1).

¹H NMR (400 MHz, CDCl₃): δ = 7.41–7.16 (m, 40 H, CH_Ph), 5.90

(m, 1 H, CH=), 5.81 (d, J_NH,2= 7.9 Hz, 1 H, NH), 5.30 (m, J_trans

= 17.2 Hz, 1 H, =CH₂), 5.22 (m, J_cis= 10.4 Hz, 1 H, =CH₂), 5.17

(br. s, 1 H, H-1_B), 5.07 (br. s, 1 H, H-1_A), 5.02–4.97 (m, 2 H, H_Bn),

4.94 (d, J_1,2= 4.0 Hz, 1 H, H-1_E), 4.90–4.86 (m, 4 H, 2H_Bn, H-1_D,

H-1_C), 4.80–4.66 (d, 8 H, H_Bn), 4.59–4.51 (m, 3 H, H_Bn), 4.38–4.32

(m, 2 H, H_All, H_Bn), 4.21 (pt, J_3,4= 9.4 Hz, 1 H, H-3_D), 4.12–4.05

(m, 6 H, H_All, H-2_B, H-3_E, H-2_A, H-3_A, H-5_C), 3.99–3.84 (m, 7 H,

H-2_C, H-5_E, H-6a_D, H-3_B, H-3_C, H-5_A, H-5_B), 3.83 (m, 1 H, H-

6b_D), 3.78 (pt, J_3,4= 9.6 Hz, 1 H, H-4_E), 3.66–3.61 (m, 2 H, H-2_E,

H-4_D), 3.57–3.52 (m, 2 H, H-4_A, H-4_B), 3.52–3.49 (m, 2 H, H-2_D,

H-6a_E), 3.43–3.35 (m, 3 H, H-6b_E, H-4_C, H-5_D), 2.02 (s, 3 H,

165.5 Hz, C-1_E), 84.1 (C-3_D), 82.4 (C-3_E), 80.2 (C-4_B), 79.8 (C-3_B),

79.4 (C-3_C), 79.3 (C-4_A), 79.2 (C-4_C), 79.0 (C-2_E), 77.8 (C-4_E), 76.5

(C-3_A), 75.8, 75.6, 75.5 (3 C, C_Bn), 75.1 (C-4_D), 75.0 (C-2_A), 75.0,

74.5, 73.4, 72.6 (5 C, C_Bn), 71.1 (C-5_D), 70.8 (2 C, C-2_C, C-5_E), 70.3

(C_All), 69.3 (C-5_B), 69.0 (C-5_C), 68.0 (C-6_E), 67.9 (C-5_A), 67.3 (C-

2_B), 62.8 (C-6_D), 57.3 (C-2_D), 23.6 (C_NAc), 18.0, 17.9, 17.8 (3 C, C-

6_A, C-6_B, C-6_C) ppm. HRMS (ESI⁺): calcd. for C₉₁H₁₀₇NO₂₃[M

+ Na]⁺1604.7131; found 1604.7180, calcd. for [M + H]⁺1582.7312;

found 1582.7300.

Propyl α-

-rhamnopyranosyl-(1Ǟ3)-α-

amido-2-deoxy-β- -glucopyranoside (6): Pentasaccharide 44

D

-Glucopyranosyl-(1Ǟ3)-α-

L-rhamnopyranosyl-(1Ǟ2)-α-

L

-rhamnopyranosyl-(1Ǟ3)-2-acet-

D

(260 mg, 164 µmol) was dissolved in EtOH (15 mL) and treated

with Pd/C (10%, 200 mg), and the suspension was stirred at room

temp. for 2 d under a hydrogen atmosphere. TLC (iPrOH/H₂O/

NH₃, 4:1:0.5 and CH₂Cl₂/MeOH, 95:5) showed that 44 had been

transformed into a more polar product. The suspension was filtered

through an Acrodisc LC (25 mm), and the filtrate was concen-

trated. Reverse phase chromatography (H₂O/CH₃CN, 100:0 Ǟ

70:30) of the residue, followed by freeze-drying, gave the target

pentasaccharide 6 (99 mg, 70%) as a white foam. Pentasaccharide

H

_NAc), 1.53 (s, 3 H, Hi_Pr), 1.43 (s, 3 H, Hi_Pr), 1.41, 1.28 (m, 6 H,

H-6_A, H-6_B), 1.23 (d, J_5,6= 6.2 Hz, 3 H, H-6_C) ppm. ¹³C NMR

(100 MHz, CDCl₃): δ = 170.4 (C_NAc), 138.6–137.6 (C_Ph), 133.8

1

6 had R_f= 0.25 (iPrOH/H₂O/NH₃, 4:1:0.5). H NMR (400 MHz,

(CH=), 129.1–127.5 (CH_Ph), 117.6 (=CH₂), 101.0 (¹J_C,H

168.4 Hz, C-1_A), 100.9 (¹J_C,H= 172.3 Hz, C-1_B), 99.8 (¹J_C,H

=

D₂O): δ = 5.15 (br. s, 1 H, H-1_B), 5.08 (d, J_1,2= 3.8 Hz, 1 H, H-

1_E), 4.98 (d, J_1,2= 1.5 Hz, 1 H, H-1_A), 4.79 (d, J_1,2= 1.2 Hz, 1 H,

H-1_C), 4.50 (d, J_1,2= 8.6 Hz, 1 H, H-1_D), 4.17 (dd, J_2,3= 2.6 Hz,

170.6 Hz, C-1_C), 99.6 (¹J_C,H= 160.5 Hz, C-1_D), 99.5 (Ci_Pr), 94.0

(¹J_C,H= 167.5 Hz, C-1_E), 82.4 (C-3_E), 80.4 (C-4_A^#), 80.3 (C-3_B^§),

79.7 (C-4_C), 79.5 (C-3_C^§), 79.3 (C-4_B^#), 79.0 (C-2_E), 77.8 (C-4_E),

76.5 (C-3_A), 76.0 (C-3_D), 75.6 (3 C, C_Bn), 75.4 (C-2_A), 75.0, 74.9,

74.4, 73.5 (4 C, C_Bn), 73.0 (C-4_D), 72.5 (C_Bn), 71.2 (C-2_C), 70.7 (C-

5_E), 70.0 (C_All), 69.1 (C-5_A*), 68.0 (C-6_E), 67.9 (C-5_B*), 67.7 (C-

5_C), 67.4 (C-2_B), 67.3 (C-5_D), 62.3 (C-6_D), 58.2 (C-2_D), 29.1 (Ci_Pr),

23.5 (C_NAc), 19.2 (Ci_Pr), 18.0, 17.8 (3 C, C-6_A, C-6_B, C-6_C) ppm.

HRMS (ESI⁺): calcd. for C₉₄H₁₁₁NO₂₃[M + Na]⁺1644.7445:

found 1644.7562, calcd. for [M + H]⁺1622.7626; found 1622.7649.

1 H, H-2_A), 4.03 (dd, J_2,3= 3.0 Hz, 1 H, H-2_B), 3.99 (m, J_4,5

=

9.8 Hz, 1 H, H-5_C), 3.94 (m, 1 H, H-5_E), 3.92–3.87 (m, 2 H, H-3_B,

H-6a_D), 3.85–3.68 (m, 11 H, H-3_A, H-2_C, H_Pr, H-2_D, H-6a_E, H-

6b_E, H-3_E, H-3_C, H-6b_D, H-5_A, H-5_B), 3.58–3.40 (m, 9 H, H-3_D,

H-4_A, H-2_E, H_Pr, H-4_C, H-4_D, H-4_B, H-4_E, H-5_D), 2.01 (s, 3 H,

H_NAc), 1.51 (sext, J = 7.2 Hz, 2 H, CH₂), 1.29–1.24 (m, 6 H, H-

6_A, H-6_B), 1.20 (d, J_5,6= 6.2 Hz, 3 H, H-6_C), 0.83 (t, J = 7.4 Hz,

3 H, CH₃) ppm. ¹³C NMR (100 MHz, D₂O): δ = 174.1 (C_NAc),

102.0 (¹J_C,H= 171.5 Hz, C-1_A), 101.3 (¹J_C,H= 173.6 Hz, C-1_C),

100.8 (¹J_C,H= 177.8 Hz, C-1_B), 100.6 (¹J_C,H= 156.2 Hz, C-1_D),

95.4 (¹J_C,H= 169.4 Hz, C-1_E), 81.7 (C-3_D), 78.3 (C-2_B), 77.3 (C-

3_C), 76.0 (C-5_D), 75.3 (C-3_A), 73.0 (C-3_E), 72.3 (C_Pr), 72.2 (C-4_B),

71.7 (2 C, C-4_C, C-5_E), 71.4 (C-4_A), 70.6 (C-2_C), 70.3 (C-2_E), 69.9

(C-3_B), 69.4 (C-4_E), 69.3 (2 C, C-5_A, C-5_B), 68.9 (C-5_C), 68.5 (C-

4_D), 66.7 (C-2_A), 60.9 (C-6_D), 60.3 (C-6_E), 55.4 (C-2_D), 22.1 (2 C,

Allyl (2,3,4,6-Tetra-O-benzyl-α-

zyl-α- -rhamnopyranosyl)-(1Ǟ2)-(3,4-di-O-benzyl-α-

pyranosyl)-(1Ǟ3)-(4-O-benzyl-α- -rhamnopyranosyl)-(1Ǟ3)-2-acet-

amido-2-deoxy-β- -glucopyranoside (44): TFA (50 % aqueous,

D

-glucopyranosyl)-(1Ǟ3)-(4-O-ben-

L

-rhamno-

L

D

4 mL) was added at 0 °C to a solution of pentasaccharide 43

(515 mg, 318 µmol) in CH₂Cl₂(10 mL), and the biphasic mixture

was stirred vigorously at room temp. for 1 h. TLC (CH₂Cl₂/MeOH,

95:5) showed the complete disappearance of the starting material

and the presence of a major more polar product. Repeated coevap-

oration with toluene and chromatography of the residue (Tol/

EtOAc, 3:7 Ǟ 1:9) provided tetraol 44 (330 mg, 66%) as a white

foam. The latter had R_f= 0.2 (CH₂Cl₂/MeOH, 95:5). ¹H NMR

(400 MHz, CDCl₃): δ = 7.42–7.13 (m, 40 H, CH_Ph), 5.90 (m, 1 H,

C

_NAc, CH₂), 20.2 (C_Ac), 16.8, 16.7 (2 C, C-6_A, C-6_B), 16.4 (C-6_C),

9.6 (CH₃) ppm. HRMS (ESI⁺): calcd. for C₃₅H₆₁NO₂₃[M + Na]⁺

886.3532; found 886.3501, calcd. for [M + H]⁺864.3713; found

864.3718.

Acknowledgments

CH=), 5.78 (d, J_NH,2= 7.2 Hz, 1 H, NH), 5.30 (m, J_trans= 17.3 Hz, This work was supported by the Agence Nationale pour la Recher-

1 H, =CH₂), 5.21 (m, J_cis= 10.4 Hz, 1 H, =CH₂), 5.17 (br. s, 1 H,

H-1_B), 5.01–4.94 (m, 4 H, H-1_D, H-1_A, 2H_Bn), 4.92 (d, J_1,2

che (ANR) (Grant NT05-1_42479), the Institut Pasteur, and the

Ministère de l’Éducation Nationale, de la Recherche et de la Techno-

=

3.7 Hz, 1 H, H-1_E), 4.89–4.86 (m, 2 H, H_Bn), 4.83 (m, J_1,2= 2.4 Hz, logie (MENRT) with a fellowship to J. B. The authors thank F.

1 H, H-1_C), 4.78–4.65 (d, 7 H, H_Bn), 4.57–4.49 (m, 3 H, H_Bn), 4.37– Bonhomme (CNRS URA 2128) for performing the NMR and

4.20 (m, 4 H, H_All, 2ϫH_Bn, H-3_D), 4.13 (m, 1 H, H_All), 4.09–4.02 mass spectra.

Eur. J. Org. Chem. 2008, 5526–5542

www.eurjoc.org

5541

J. Boutet, L. A. Mulard

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Received: July 13, 2008

Published Online: October 13, 2008

5542

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Article Doi

Synthesis of two tetra- and four pentasaccharide fragments of Shigella flexneri serotypes 3a and X O-antigens from a common tetrasaccharide intermediate

DOI: 10.1002/ejoc.200800693

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