Synthesis of new glycopolymers containing β-d-mannopyranose, and c-2-substituted β-d-mannopyranose residues as a new class of inhibitor

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SYNTHESIS OF NEW GLYCOPOLYMERS CONTAINING

β-D-MANNOPYRANOSE, AND C-2-SUBSTITUTED β-

D-MANNOPYRANOSE RESIDUES AS A NEW CLASS OF

INHIBITOR

Shoji Akai ^a, Yasuhiro Kajihara ^b, Yasuhiko Nagashima ^a, Masugu Kamei ^c, Junko Arai ^c,

Masami Bito ^c& Ken-ichi Sato ^d

^aLaboratory of Organic Chemistry, Faculty of Engineering , Kanagawa University ,

3-27-1, Rokkakubashi, Kanagawa-ku, Yokohama, 221-8686, Japan

^bDepartment of System Functions, Faculty of Science , Yokohama City University , 22-2,

Seto, Kanazawa-ku, Yokohama, 236-0027, Japan

^cSugiyama Chemical & Industrial Laboratory , 11, Kagetori-cho, Totsuka-ku, Yokohama,

245-0046, Japan

^dLaboratory of Organic Chemistry, Faculty of Engineering , Kanagawa University ,

3-27-1, Rokkakubashi, Kanagawa-ku, Yokohama, 221-8686, Japan

Published online: 16 Aug 2006.

To cite this article: Shoji Akai , Yasuhiro Kajihara , Yasuhiko Nagashima , Masugu Kamei , Junko Arai , Masami Bito &

Ken-ichi Sato (2001) SYNTHESIS OF NEW GLYCOPOLYMERS CONTAINING β-D-MANNOPYRANOSE, AND C-2-SUBSTITUTED β-

D-MANNOPYRANOSE RESIDUES AS A NEW CLASS OF INHIBITOR, Journal of Carbohydrate Chemistry, 20:2, 121-143, DOI:

10.1081/CAR-100103953

To link to this article: http://dx.doi.org/10.1081/CAR-100103953

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J. CARBOHYDRATE CHEMISTRY, 20(2), 121–143 (2001)

SYNTHESIS OF NEW GLYCOPOLYMERS

CONTAINING ꢀ-D-MANNOPYRANOSE, AND

C-2-SUBSTITUTED ꢀ-D-MANNOPYRANOSE

RESIDUES AS A NEW CLASS OF INHIBITOR

Shoji Akai,¹Yasuhiro Kajihara,²Yasuhiko Nagashima,¹

Masugu Kamei,³Junko Arai,³Masami Bito,³and Ken-ichi Sato^1,*

¹Laboratory of Organic Chemistry, Faculty of Engineering,

Kanagawa University, 3-27-1, Rokkakubashi, Kanagawa-ku,

Yokohama, 221-8686 Japan

²Department of System Functions, Faculty of Science,

Yokohama City University, 22-2, Seto, Kanazawa-ku,

Yokohama, 236-0027 Japan,

³Sugiyama Chemical & Industrial Laboratory, 11, Kagetori-cho,

Totsuka-ku, Yokohama, 245-0046 Japan

ABSTRACT

New styryl monomers containing ꢀ-D-mannopyranose, 2-acetamido-2-deoxy-

ꢀ-D-mannopyranose, 2-deoxy-2-fluoro-ꢀ-D-mannopyranose, and 2-deoxy-ꢀ-

D-arabino-hexopyranose on their side chains, were efficiently synthesized as a

new class of a potent inhibitor resistant to exo-ꢁ-mannosidase digestion. The

binding affinity of the carbohydrate polymers obtained from those mannopy-

ranosyl styryl monomers by radical polymerization with Concanavalin A were

evaluated. A binding assay indicated that the multivalency effect and the affin-

ity enhancement attained by modification at the C-2 position of the ꢀ-D-

mannopyranoside residue resulted in the ꢀ-D-mannopyranosyl polymer which

has the same affinity as that of the ꢁ-D-mannopyranosyl polymer.

*Corresponding author.

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INTRODUCTION

Concanavalin A (Con A) binds ꢁ-D-mannopyranoside in preference to other

mono-carbohydrate ligands and this specificity has been extensively investi-

gated.¹The 3-, 4-, and 6-hydroxyl groups of D-mannopyranoside are essential for

Con A binding. On the other hand, the Con A binding affinity toward ꢀ-D-

mannopyranoside is only one-fortieth that of ꢁ-D-mannopyranoside¹due to steric

hindrance of the glycosyl bond in the binding site.¹In addition, X-ray crystallo-

graphic analysis showed that the oxygen atom at the anomeric position and the

2-hydroxyl group of ꢁ-D-mannopyranoside is positioned out of the binding site

of Con A.²This result suggests that the 2-hydroxyl group is not specifically rec-

ognized.

In order to synthesize an effective inhibitor against carbohydrate-processing

enzymes, for use as in in vivo assays, resistance toward exo-glycosidase digestion

is essential. With this goal in mind, several inhibitors such as S- or C-glycosyl ana-

logues have been synthesized by other researchers.^3,4In our work, we planned to

synthesize a new class of inhibitor based on a ꢀ-D-mannopyranoside scaffold,

which can resist exo-ꢁ-D-mannosidase, which is relatively abundant compared

with the low levels of ꢀ-D-mannosidase present in the in vivo assay environment.

However, the binding affinity of ꢀ-D-mannopyranoside is significantly less than

that of the ꢁ-isomer, as mentioned above. In order to enhance the low binding

affinity of a ꢀ-D-mannopyranoside scaffold, several methods were considered.

Recently, two approaches for enhancement of the binding affinity of carbo-

hydrate ligands have been reported. One is the use of a multivalent glycoconjugate

to take advantage of the multivalency effect.^5–16The other approach is to modify

the carbohydrate ligand with an enhancer¹⁷for binding affinity. Specifically, one

of the hydroxyl groups positioned away from the binding site of the receptor, is

modified with an enhancer. We thought that these two approaches could be com-

bined effectively to enhance the binding affinity of carbohydrate ligands to recep-

tors. However, such a combined approach had not been reported. Therefore, we set

out to determine whether a combined multivalency effect and enhancer modifica-

tion would result in ꢀ-D-mannopyranoside possessing sufficient binding affinity

toward Con A. We expected the substitution of the 2-hydroxyl group of ꢀ-D-

mannopyranoside with an enhancer to result in increased binding affinity. In this

work, we examined substitution of the 2-hydroxyl with hydrogen (deoxy),^18,19flu-

oro,^18,19and acetamide groups, because hydrogen atoms are known to enhance

binding affinity through hydrophobic interactions and fluorine acts as a proton

acceptor for hydrogen bonds. The acetamide group is thought to act as both a pro-

ton donor and a proton acceptor in addition to participating in hydrophobic

interactions.

In this paper, we describe the synthesis and the Con A binding ability of

several styryl polymers containing ꢀ-D-mannopyranoside analogues containing

binding enhancers to illustrate this approach for the construction of novel in-

hibitors.

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SYNTHESIS OF NEW GLYCOPOLYMERS

123

RESULTS AND DISCUSSION

Synthesis

For the synthesis of glycopolymers, we selected a styryl derivative as the

monomer. Further, 3-(p-hydroxyphenyl)propionic acid was selected as the spacer,

since an aromatic ring at the anomeric position of D-mannopyranoside increases

the binding affinity over that of the methyl glycoside.¹In addition, the propionic

acid residue was used to distance the polymer backbone from the carbohydrate

residue and thus enable efficient binding with Con A.

We started by synthesizing the glycopolymers 6 containing the spacer-armed

ꢁ-D-mannopyranoside as the standard compound for the Con A binding assay. The

ꢁ-D-mannopyranoside derivative 2 was prepared by glycosylation of 1,2,3,4,6-

penta-O-acetyl-D-mannopyranose with benzyl 3-(p-hydroxyphenyl)propionate in

83% yield, and then reduced in the presence of 10% Pd(OH)₂-C/H₂in EtOH to

give the corresponding carboxylic acid derivative 3 in 97% yield. Condensation of

3 with p-vinylbenzylamine¹⁶by the use of 1-ethyl-3-(3-dimethylaminopropyl)car-

bodiimide (WSC) in dichloromethane gave the corresponding amide derivative 4

in 73% yield. Subsequently, O-deacetylation of 4 with NaOMe/MeOH gave p-[2-

(N-p-vinylbenzyl)carbamoyl]ethyl]phenyl ꢁ-D-mannopyranoside 5, in 97% yield,

as a glycomonomer containing the ꢁ-D-mannopyranoside residue on its side chain.

Finally, the ꢁ-D-mannopyranosyl polymer 6 was obtained via radical polymeriza-

tion of 5 by treatment with 2,2ꢂ-azobisisobutyronitrile (AIBN), in 98% yield

(Scheme 1).

For the synthesis of the ꢀ-D-mannopyranoside derivatives, we employed a

method developed previously in our laboratory to form the ꢀ-D-mannopyranosyl

linkage (Scheme 2).²⁰In this way, we synthesized glycomonomers containing the

Scheme 1.

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AKAI ET AL.

Scheme 2.

spacer-armed ꢀ-D-mannopyranoside derivative 12, the 2-acetamido-2-deoxy-ꢀ-D-

mannopyranoside derivative 15, the 2-deoxy-ꢀ-D-arabino-hexopyranoside deriva-

tive 16, and the 2-deoxy-2-fluoro-ꢀ-D-mannopyranoside derivative 17 starting

from the ꢀ-D-galactopyranoside derivative 10 (Schemes 2 and 3). Compound 10

was synthesized as follows. Glycosylation of 1,2,3,4,6-penta-O-acetyl-ꢀ-D-galac-

topyranose 7 with benzyl 3-(p-hydroxyphenyl)propionate in the presence of

tin(IV) chloride gave the corresponding ꢀ-D-galactopyranoside 8 in 59% yield,

which was then treated with NaOBn/BnOH in dichloromethane to give p-[2-(ben-

Scheme 3.

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SYNTHESIS OF NEW GLYCOPOLYMERS

125

zyloxycarbonyl)ethyl]phenyl ꢀ-D-galactopyranoside 9 in 74% yield. Selective

stannylation of 9 with bis(tributyltin) oxide and subsequent pivaloylation were per-

formed in 66% yield (2 steps). The resulting pivaloate 10 was then treated with tri-

fluoromethanesulfonic anhydride in pyridine at ꢃ19°C to give the 2,4-bistriflate

derivative 11 in quantitative yield. Simultaneous nucleophilic substitution reac-

tions at the C-2 and C-4 positions of 11 with cesium acetate (3.0 mol equiv) af-

forded the ꢀ-D-mannopyranoside derivative 12 in 87% yield. The structure of 12

1

was identified by H NMR (J_3,4ꢄ 9.7 Hz, J_4,5ꢄ 9.7 Hz, J_1,2ꢄ 1.2 Hz and J_2,3

ꢄ 3.0 Hz). When a small amount of cesium acetate (1.2 mol equiv) was used, the

reaction afforded the 4-O-acetyl glucopyranoside 13 exclusively (92%) (Scheme

2). This ꢀ-D-glucopyranoside derivative (2-triflate) 13 was treated with n-Bu₄NN₃,

n-Bu₄NBH₄,^20,21or n-Bu₄NF,^20,22under ultrasonication conditions to afford the

corresponding manno-type 2-azido 14, 2-deoxy 16, and 2-fluoro 17 derivatives, re-

spectively, in moderate yields (Scheme 3). Catalytic reduction of the azido group

of 14 and subsequent acetylation gave the desired 2-acetamido-ꢀ-D-mannopyra-

noside derivative 15 in 53% yield as well as unreacted 14 (42%). ꢀ-D-Mannopyra-

nosyl derivatives 12, 15, 16, and 17 thus obtained were then hydrogenolyzed to

give the corresponding carboxylic acid derivatives 18 (95%), 19 (91%), 20 (92%),

and 21 (90%), respectively. Condensation of 18, 19, 20, and 21 with p-vinylben-

zylamine¹⁶led to the corresponding amide derivatives 22, 23, 24, and 25 in 67, 81,

76, and 61% yields, respectively. O-Deacylation of 22, 23, 24, and 25 with

NaOMe/MeOH, was performed in near quantitative yield to afford the desired

p-[3-(N-p-vinylbenzyl)carbamoyl]ethyl]phenyl glycomonomers of ꢀ-D-mannopy-

ranoside 26, 2-deoxy-2-acetamido-ꢀ-D-mannopyranoside 27, 2-deoxy-ꢀ-D-

arabino-hexopyranoside 28, and 2-deoxy-2-fluoro-ꢀ-D-mannopyranoside 29,

respectively. A polymerization reaction with 2,2ꢂ-azobisisobutyronitrile (AIBN) in

dimethylsulfoxide was performed to give the corresponding glycopolymers 30, 31,

32 and 33 containing ꢀ-D-mannopyranoside (97% yield), 2-acetamido-2-deoxy-ꢀ-

D-mannopyranoside (84% yield), 2-deoxy-2-fluoro-ꢀ-D-mannopyranoside (86%

yield), 2-deoxy-ꢀ-D-arabino-hexopyranoside (56% yield) residues, respectively

(Scheme 4). The averaged molecular weights and reduced viscosities were ana-

lyzed by gel permeation chromatography (GPC). The physical properties of the

synthesized glycopolymers are shown in Table 1.

Con A Binding Assay

Binding assays of the synthetic glycopolymers toward Con A was performed

using the enzyme linked lectin assay (ELLA)^23–25method, which is a standard as-

say in the field. Soybean lectin with high mannose-type sugar chains was coated

on a 96-well test plate and then used for the ELLA. The results are shown in Table

2.

The inhibition abilities of methyl ꢁ-D-mannopyranoside 35 and methyl ꢁ-D-

glucopyranoside 36 show the same relative binding abilities in our assay system as

in the published data.²⁴The binding affinity of methyl ꢁ-D-glucopyranoside 36

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AKAI ET AL.

Table 1. Physical Properties of Glycopolymers

Polymers

Mn^aꢅ 10^ꢃ4

Mw / Mn^a

ꢆ_red[dL/g]^b

6

30

31

32

33

6.3

5.1

2.3

5.3

7.6

1.90

1.45

1.65

1.63

1.53

0.47

0.12

0.10

0.18

0.33

^aDetermined by GPC based on polystyrene;

^bMeasured at 30 °C in DMSO.

Table 2. Inhibition Ability of Glycopolymers

Number of

Monomer

Relative

Potency

Polymers

IC₅₀[M]

Monovalency-IC₅₀[mM]

6

30

31

1.51 ꢅ 10²

1.22 ꢅ 10²

50.1

2.0 ꢅ 10^ꢃ2

3.1 ꢅ 10^ꢃ1

—

3.0

37

—

5.6 ꢅ 10²

45

—

2.1

32

1.32 ꢅ 10²

1.81 ꢅ 10²

1.00

6.0

8.0 ꢅ 10²

7.2

33

4.0 ꢅ 10^ꢃ2

1.7 ꢅ 10³

2.5 ꢅ 10³

2.4 ꢅ 10²

1.0

35^a

36^b

1.7 ꢅ 10³

2.5 ꢅ 10³

1.00

6.9 ꢅ 10^ꢃ1

^aMethyl ꢁ-D-mannopyranoside: 35; ^bMethyl ꢁ-D-glucopyranoside: 36.

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127

was measured to be 70% of that of ꢁ-D-mannopyranoside 35. In polymers, the in-

hibition ability of 2-deoxy-ꢀ-D-mannosyl polymer 32 is shown to be only 2.1 times

stronger than that of ꢁ-D-mannopyranoside 35 despite the multivalency. Also, 2-

deoxy-2-acetamido-ꢀ-D-mannosyl polymer 31 showed no inhibitory activity. For

a natural-type mannopyranosyl polymer, ꢁ-D-mannopyranosyl polymer 6 dis-

played potent inhibition activity (3.0 ꢇM/residue). The binding affinity of the ꢀ-

D-mannopyranosyl polymer 30 was one-twelfth of that of the ꢁ-D-mannopyranosyl

polymer 6 due to the multivalency effect. Further, the inhibition potency was dra-

matically improved by the fluoro substituent of 33 resulting in a level comparable

to that of the ꢁ-D-mannopyranosyl polymer 6.

Con A recognizes ꢁ-D-mannopyranoside in strong preference to other mono-

carbohydrate ligands. When ꢀ-D-mannopyranoside is used as a ligand, the binding

affinity is one-fortieth of that of ꢁ-D-mannopyranoside.¹This tendency was also

observed with the carbohydrate polymers 6 and 30. The binding ability of the

ꢀ-D-mannopyranosyl polymer 30 is one-twelfth of that of the corresponding ꢁ-D-

mannopyranosyl polymer 6, but the binding ability of ꢀ-D-mannopyranosyl poly-

mer 30 is 45 times larger than that of methyl ꢁ-D-mannopyranoside 36. These

enhanced binding affinities almost certainly result from the multivalency effect.

The hydrogen (deoxy) and acetamide groups at C-2 did not result in a notable en-

hancement in binding affinity. In contrast, the fluorine atom at C-2 enhanced the

binding affinity of the ꢀ-D-mannopyranosyl polymer to the same level as that of

the ꢁ-D-mannopyranosyl polymer. From this result we speculate that the fluorine

atom at C-2 forms a hydrogen bond with an amino acid at the entrance of the Con

A binding site, resulting in enhanced binding affinity. Recently, H. Tanaka and

co-workers synthesized a glycoconjugate polymer carrying a 3,6-branched ꢁ-D-

mannopyranosyl trisaccharide²⁶on its scaffold.²⁷Although it would seem that us-

ing a natural-type branched oligosaccharide residue would be most effective for

increasing the binding ability, it is also important to consider the special properties

of synthetic derivatives as inhibitors for use with in vivo assays, both for binding

affinity and for resistance to exo-glycosidase digestion.

In conclusion, we have succeeded in the synthesis of new styryl polymers

containing ꢀ-D-mannopyranose derivatives and found them to be potent inhibitors

of exo-ꢁ-mannosidase digestion. We believe this to be the first report of such

polymers. The results of Con A binding affinity studies show that some of these

novel glycopolymers display impressive properties. We feel that the new approach

described herein, which combines the multivalency effect and enhancer tech-

niques, constitutes an exciting advance for the synthesis of highly potent inhibitors

resistant to exo-glycosidase digestion.

EXPERIMENTAL

General Methods. All melting points were uncorrected. The solutions

were concentrated under reduced pressure at a bath temperature not exceeding

45°C. The optical rotations were measured in a 0.5 dm tube with JASCO DIP-140

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AKAI ET AL.

polarimeter in chloroform unless otherwise stated. ¹H NMR spectra were recorded

with a JEOL EX-90, JEOL FX-200, JEOL EX-270, or JEOL A-500 spectrometer.

IR spectra were recorded with a Hitachi 270-30 spectrometer. Elemental analyses

were performed on a Perkin-Elmer 2400 II elemental analyzer. The chemical

shifts, coupling constants, and IR frequencies were recorded in ꢈ, Hz, and cm^ꢃ1

units, respectively. Column chromatography was performed on silica gel (Silica

gel 60, 70-230 mesh, Merck). Thin-layer chromatography (TLC) on silica gel (Sil-

ica gel 60 F₂₅₄, Merck) was used to monitor the reactions and to certify the reac-

tion products. The molecular weights of the glycopolymers were estimated by gel

permeation chromatography (GPC) with the use of a TOSO Model HLC-8020

GPC equipped with a refractive index detector using TSK gel columns (Tosoh Co.

Ltd.) (eluent: N,N-dimethylformamide, calibrated with narrow molecular weight

polystyrene standards).

p-[2-(Benzyloxycarbonyl)ethyl]phenyl

2,3,4,6-Tetra-O-acetyl-ꢁ-D-

mannopyranoside (2). To a solution of 1,2,3,4,6-penta-O-acetyl-ꢁ,ꢀ-D-

mannnopyranose (1) (558 mg, 1.6 mmol), benzyl 3-(p-hydroxyphenyl)propionate

(34) (665 mg, 2.6 mmol), and molecular sieves 4A in dry dichloromethane (20

mL), trimethylsilyl trifluoromethanesulfonate (0.31 mL, 1.6 mmol) was added

dropwise under argon at 0°C. The reaction mixture was stirred at room tempera-

ture for 12 h until the disappearance of 1 on TLC with hexane/ethyl acetate (3/1

v/v). Triethylamine (1 mL) was added to the reaction mixture at 0°C, then molec-

ular sieves were filtered off. The filtrate was poured into a saturated aq NaHCO₃

solution, extracted with ethyl acetate, washed with brine and water, dried over an-

hydrous magnesium sulfate, filtered, and concentrated to give a residue. The re-

maining residue was purified on a column of silica gel with hexane/ethyl acetate

(3/1 v/v) to give 2 (692 mg, 83%): [ꢁ]_D²⁵ꢉ58.2° (c 2.1, CHCl₃); IR (cm^ꢃ1) 1743

1

(CꢄO); H NMR (FX-200, CDCl₃) ꢈ 7.37–7.29 (m, 5H, -CH₂Ph, J ꢄ 8.8 Hz),

7.12, 6.99 (each d, 4H, phenylene), 5.56 (dd, 1H, H-3, J_3,4ꢄ 9.9 Hz), 5.47 (dd, 1H,

H-1, J_1,2ꢄ 1.8 Hz), 5.43 (dd, 1H, H-2, J_2,3ꢄ 3.6 Hz), 5.37 (t, 1H, H-4, J_4,5ꢄ 10.1

Hz), 5.10 (s, 2H, -CH₂Ph), 4.27 (dd, 1H, H-6ꢂ, J_5,6ꢂꢄ 5.1 Hz), 4.10 (ddd, 1H, H-

5, J_5,6ꢄ 2.4 Hz), 4.05 (dd, 1H, H-6, J_6,6ꢂꢄ 12.2 Hz), 2.93 (t, 2H, -CH₂CH₂CO-,

J ꢄ 8.0 Hz), 2.65 (t, 2H, -CH₂CH₂CO-), 2.19, 2.05, 2.03, 2.02 (each s, 3H ꢅ 4,

OAc).

Anal. Calcd for C₃₀H₃₄O₁₂: C, 61.43; H, 5.84. Found: C, 61.41; H, 5.88.

p-(2-Carboxyethyl)phenyl 2,3,4,6-Tetra-O-acetyl-ꢁ-D-mannopyranoside

(3). p-[2-(Benzyloxycarbonyl)ethyl]phenyl glycoside 2 (1.60 g, 2.7 mmol) was

hydrogenolyzed in ethanol (25 mL) in the presence of a catalytic amount of 10%

Pd(OH)₂-C/H₂for 2 h until the disappearance of 2 on TLC with hexane/ethyl ac-

etate (3/1 v/v). The solution was then filtered through a pad of celite and concen-

25

trated to afford 3 (1.34 g, 97% yield): [ꢁ]_Dꢉ67.4° (c 0.8, CHCl₃); IR (cm^ꢃ1

3192 (OH), 1748 (CꢄO); H NMR (FX-200, CDCl₃) ꢈ 7.13, 7.00 (each d, 4H,

phenylene, J ꢄ 8.1 Hz), 6.78 (br s, 1H, COOH), 5.55 (dd, 1H, H-3, J_3,4ꢄ 10.0 Hz),

5.48 (dd, 1H, H-1, J_1,2ꢄ 1.9 Hz), 5.43 (dd, 1H, H-2, J_2,3ꢄ 3,4 Hz), 5.36 (dd, 1H,

)

1

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129

H-4, J_4,5ꢄ 10.0 Hz), 4.28 (dd, 1H, H-6ꢂ, J_5,6ꢂꢄ 5.1 Hz, J_6,6ꢂꢄ 12.0 Hz), 4.13–4.04

(m, 2H, H-5 and H-6), 2.89 (t, 2H, -CH₂CH₂CO-, J ꢄ 7.8 Hz), 2.61 (t, 2H, -

CH₂CH₂CO-), 2.20, 2.05, 2.03, 2.03 (each s, 3H ꢅ 4, OAc).

Anal. Calcd for C₂₃H₂₈O₁₂: C, 55.65; H, 5.68. Found: C, 55.45; H, 5.72.

p-[2-[N-(p-Vinylbenzyl)carbamoyl]ethyl]phenyl 2,3,4,6-Tetra-O-acetyl-

ꢁ-D-mannopyranoside (4). To a solution of p-(2-carboxyethyl)phenyl glyco-

side 3 (1.27 g, 2.50 mmol) and 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide

(WSC) (585 mg, 3.05 mmol) in dichloromethane (100 mL), a solution of p-vinyl-

benzylamine¹⁶(378 mg, 2.82 mmol) in dichloromethane (40 mL) was added

dropwise at 0°C, and stirred for 1 h until the disappearance of 3 on TLC with

hexane/ethyl acetate (1/1 v/v). The reaction mixture was diluted with di-

chloromethane, washed with 10 % aq citric acid solution, brine and water, dried

over anhydrous magnesium sulfate, filtered, and concentrated to give a residue.

The remaining residue was purified on a column of silica gel with hexane/ethyl ac-

25

etate (3/5 v/v) to afford 4 (1.14 g, 73% yield): [ꢁ]_Dꢉ66.0° (c 0.7, CHCl₃); IR

(cm^ꢃ1) 1758 (CꢄO), 1650 (CꢄC); ¹H NMR (A-500, CDCl₃) ꢈ 7.34, 7.12 (each d,

4H, styryl phenylene, J ꢄ 7.9 Hz), 7.12, 6.99 (each d, 4H, phenylene, J ꢄ 8.5 Hz),

6.69 (dd, 1H, -Ph-CHꢄCHaHb, J_transꢄ 17.6 Hz, J_cisꢄ 11.2 Hz), 5.73 (dd, 1H, Ha,

J_gemꢄ 0.7 Hz), 5.71 (t, 1H, -CONH-), 5.56 (dd, 1H, H-3, J_3,4ꢄ 9.9 Hz), 5.47 (dd,

1H, H-1, J_1,2ꢄ 1.8 Hz), 5.43 (1H, dd, H-2, J_2,3ꢄ 3.6 Hz), 5.37 (1H, dd, H-4, J_4,5

ꢄ 10.1 Hz), 5.23 (1H, dd, Hb), 4.37 (2H, d, -NHCH₂-, J_NH,CH2ꢄ 5.8 Hz), 4.27

(1H, dd, H-6ꢂ, J_5,6ꢂꢄ 5.1 Hz), 4.10 (1H, ddd, H-5, J_5,6ꢄ 2.4 Hz), 4.05 (dd, 1H, H-

6, J_6,6ꢂꢄ 12.2 Hz), 2.94 (t, 2H, -CH₂CH₂CO-, J ꢄ 7.8 Hz), 2.47 (t, 2H,

-CH₂CH₂CO-), 2.19, 2.05, 2.03, 2.02 (each s, 3H ꢅ 4, OAc).

Anal. Calcd for C₃₂H₃₇O₁₁N: C, 62.84; H, 6.10; N, 2.15. Found: C, 62.42; H,

6.55; N, 2.26.

p-[2-[N-(p-Vinylbenzyl)carbamoyl]ethyl]phenyl ꢁ-D-Mannopyranoside

(5). To a solution of p-[2-[N-(p-vinylbenzyl)carbamoyl]ethyl]phenyl glycoside 4

(1.14 g, 1.86 mmol) in methanol (25 mL), sodium methoxide (1.9M methanol so-

lution) was carefully added at pH 10, stirred for 2 h until the disappearance of 4 on

TLC with hexane/ethyl acetate (1/2 v/v). The reaction mixture was neutralized

with Dowex 50W-X8 (H^ꢉform) ion-exchange resin, filtered, and concentrated to

give pure styryl monomer containing carbohydrate residue 5 (756 mg, 97% yield):

mp 171–174°C (methanol); [ꢁ]_D²⁵ꢉ83.6° (c 0.7, CHCl₃); IR (cm^ꢃ1) 3284 (OH),

1640 (CꢄO, CꢄC); ¹H NMR (A-500, d₆-DMSO) ꢈ 8.35 (t, 1H, NH), 7.33, 6.98

(each d, 4H, styryl phenylene, J ꢄ 8.2 Hz), 7.12, 7.11 (each d, 4H, phenylene,

J ꢄ 8.6 Hz), 6.68 (dd, 1H, -Ph-CHꢄCHaHb, J_transꢄ 17.4 Hz, J_cisꢄ 10.8 Hz),

5.76, 5.73 (dd, 1H, Ha, J_gemꢄ 0.9 Hz), 5.35 (d, 1H, H-1, J_1,2ꢄ 1.9 Hz), 5.21 (dd,

1H, Hb), 4.26 (d, 2H, -NHCH₂-, J_NH,CH2ꢄ 5.5 Hz), 3.89 (d, 1H, H-2, J_2,3ꢄ 3.3

Hz), 3.75 (dd, 1H, H-3, J_3,4ꢄ 9.2 Hz), 3.63–3.57 (m, 3H, H-4, H-6, and H-6ꢂ), 3.49

(ddd, 1H, H-5, J_4,5ꢄ 9.5 Hz, J_5,6ꢄ 2.7 Hz, J_5,6ꢂꢄ 5.2 Hz), 3.39 (s, 4H, OH), 2.83

(t, 2H, -CH₂CH₂CO-, J ꢄ 7.8 Hz), 2.45 (t, 2H, -CH₂CH₂CO-); ¹³C NMR (A-500,

d₆-DMSO) ꢈ 171.32 (CꢄO), 138.87, 135.54, 134.46, 128.95, 127.20, 125.71 (Ph),

ORDER

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130

AKAI ET AL.

136.12 (CHꢄCH₂), 113.33 (CHꢄCH₂), 98.95 (C-1), 74.14 (C-2), 70.61 (C-3),

69.96 (C-4), 66.67 (C-5), 60.90 (C-6), 41.69, 37.11, 30.29 (CH₂).

Anal. Calcd for C₂₄H₂₉O₇₆N: C, 65.00; H, 6.59; N, 3.16. Found: C, 64.82; H,

6.30; N, 2.70.

Poly[p-[2-[N-(p-vinylbenzyl)carbamoyl]ethyl]phenyl ꢁ-D-mannopyra-

noside] (6). To a solution of 5 (726 mg, 1.64 mmol) in DMSO (2.0 mL), 2,2ꢂ-

azobisisobutyronitrile (4.0 mg, 0.02 mmol) was added. The mixture was set in a

sealed tube, degassed with argon, and kept at 65°C in oil bath for 24 h. The reac-

tion mixture was poured into stirred acetone, and the precipitate was filtered using

a glass filter to give 6 (710 mg, 98% yield): IR (cm^ꢃ1) 3408 (OH), 1646 (CꢄO);

¹³C NMR (A-500, d₆-DMSO) ꢈ 171.32 (CꢄO), 128.99, 116.65 (Ph), 126.52 (-CH-

CH₂-), 99.00 (C-1), 74.58 (C-2), 70.66 (C-3), 70.09 (C-4), 66.69 (C-5), 60.99

(C-6), 41.72, 40.41, 37.19, 30.33 (CH₂).

p-[2-(Benzyloxycarbonyl)ethyl]phenyl 2,3,4,6-Tetra-O-acetyl-ꢀ-D-galac-

topyranoside (8). To a mixture of 7 (15.0 g, 384 mmol) and 34 (10.8 g, 422 mmol)

in dry CH₂Cl₂(300 mL), anhydrous tin(IV) chloride (5.0 mL, 422 mmol) was added

dropwise under argon at ꢃ19°C. The reaction mixture was kept with stirring for 1

h until the disappearance of 7 on TLC with hexane/ethyl acetate (3/1 v/v). The re-

action mixture was poured into a saturated aq NaHCO₃solution with activated

potassium fluoride (5.0 g), and stirred for 12 h. The above mixture was filtered and

extracted with CHCl₃. The combined organic solution was washed with brine and

water, dried over anhydrous magnesium sulfate, filtered, and concentrated to give

a residue. The remaining residue was purified on a column of silica gel with hex-

ane/ethyl acetate (3/1 v/v) to afford 8 (13.8 g, 59% yield): mp 108–109°C (ethanol-

hexane); [ꢁ]_D²⁵ꢃ13.4° (c 0.9, CHCl₃); IR (cm^ꢃ1) 1752 (CꢄO); ¹H NMR (FX-200,

CDCl₃) ꢈ 7.37–7.26 (m, 4H, -CH₂Ph), 7.11, 6.99 (each d, 4H, phenylene, J ꢄ 8.8

Hz), 5.48 (dd, 1H, H-2, J_2,3ꢄ 10.5 Hz), 5.45 (dd, 1H, H-4, J_4,5ꢄ 0.9 Hz), 5.10 (dd,

1H, H-3, J_3,4ꢄ 3.4 Hz), 5.10 (s, 2H, -CH₂Ph), 5.00 (d, 1H, H-1, J_1,2ꢄ 7.8 Hz),

4.25–4.04 (m, 3H, H-5, H-6, and H-6ꢂ), 2.93 (t, 2H, -CH₂CH₂CO-, J ꢄ 7.8 Hz), 2.65

(t, 2H, -CH₂CH₂CO-), 2.18, 2.07, 2.06, 2.01 (each s, 3H ꢅ 4, OAc).

Anal. Calcd for C₃₀H₃₄O₁₂: C, 61.43; H, 5.84. Found: C, 61.41; H, 5.88.

p-[2-(Benzyloxycarbonyl)ethyl]phenyl ꢀ-D-Galactopyranoside (9). To

a solution of 8 (200 mg, 0.34 mmol) in CH₂Cl₂(50 mL), 1M NaOBn in BnOH was

carefully added at pH 11 at 0°C, and stirred for 1 h until the disappearance of 9 on

TLC with hexane/ethyl acetate (3/1 v/v). The reaction mixture was neutralized

with Dowex 50W-X8 (H^ꢉform) ion-exchange resin, filtered, and concentrated to

give a residue. The remaining residue was purified on a column of silica gel with

25

CHCl₃/MeOH (8/1 v/v) to afford 9 (120 mg, 74% yield): [ꢁ]_Dꢃ25.4° (c 0.9,

MeOH); mp 86–87.5°C (methanol-ether); IR (cm^ꢃ1) 3369 (OH), 1730 (CꢄO); ¹H

NMR (A-500, CD₃OD) ꢈ 7.24–7.14 (m, 5H, -CH₂Ph), 7.11, 7.02 (each d, 4H,

phenylene, J ꢄ 8.8 Hz), 5.10 (s, 2H, -CH₂Ph), 4.84 (d, 1H, H-1, J_1,2ꢄ 7.6 Hz),

ORDER

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SYNTHESIS OF NEW GLYCOPOLYMERS

131

3.93 (dd, 1H, H-4, J_4,5ꢄ 1.0 Hz), 3.82 (dd, 1H, H-2, J_2,3ꢄ 9.8 Hz), 3.81 (dd, 1H,

H-6ꢂ. J_6,6ꢂꢄ 11.3 Hz), 3.77 (dd, 1H, J_5,6ꢄ 6.7 Hz), 3.69 (ddd, 1H, H-5, J_5,6ꢂꢄ 5.2

Hz), 3.60 (dd, 1H, H-3, J_3,4ꢄ 3.7 Hz), 2.90 (t, 2H, -CH₂CH₂CO-, J ꢄ 7.6 Hz), 2.66

(t, 2H, -CH₂CH₂CO-).

Anal. Calcd for C₂₂H₂₆O₈: C, 63.15; H, 6.26. Found: C, 63.35; H, 6.26.

p-[2-(Benzyloxycarbonyl)ethyl]phenyl 3,6-Di-O-pivaloyl-ꢀ-D-galactopy-

ranoside (10). A solution of 9 (300 mg, 0.72 mmol) and bis(tributyltin) oxide

(1.31 g, 2.2 mmol) in toluene was refluxed for 2 h with a Dean-Stark apparatus,

then pivaloyl chloride (2.5 mL, 2.09 mmol) was added at 0°C and stirred for 72 h

at rt until the disappearance of 9 on TLC with chloroform/methanol (8/1 v/v). The

mixture was diluted with ethyl acetate (50 mL), treated with a saturated aq

NaHCO₃solution and activated potassium fluoride, filtered, and separated. The

aqueous solution was extracted with ethyl acetate. The combined organic solution

was washed with brine and water, dried over anhydrous magnesium sulfate, fil-

tered, and concentrated to give a residue. The remaining residue was purified on a

column of silica gel with hexane/ethyl acetate (2/1 v/v) to afford 10 (278 mg, 66%

yield): mp 104–105°C (ethanol-hexane); [ꢁ]_D²⁵ꢉ17.4° (c 0.5, CHCl₃); IR (cm^ꢃ1

)

3548 (OH), 1724 (CꢄO); ¹H NMR (A-500, CDCl₃) ꢈ 7.37–7.28 (m, 5H, -CH₂Ph),

7.08, 6.97 (each d, 4H, phenylene, J ꢄ 8.5 Hz), 5.10 (s, 2H, -CH₂Ph), 4.93 (dd, 1H,

H-3, J_3,4ꢄ 10.1 Hz), 4.88 (d, 1H, H-1, J_1,2ꢄ 7.7 Hz), 4.36 (dd, 1H, H-6, J_6,6ꢂ

ꢄ 11.6 Hz), 4.29 (dd, 1H, H-6ꢂ), 4.12 (ddd, 1H, H-2, J_2,3ꢄ 3.4 Hz), 4.04 (dd, 1H,

H-4), 3.89 (dd, 1H, H-5, J_5,6ꢄ 5.2 Hz, J_5,6ꢂꢄ 7.4 Hz), 2.91 (t, 2H, -CH₂CH₂CO,

J ꢄ 7.7 Hz), 2.64 (t, 2H, -CH₂CH₂CO-), 2.46 (d, 1H, OH, J_2,OHꢄ 3.3 Hz), 2.21

(d, 1H, OH, J_4,OHꢄ 4.9 Hz), 1.27, 1.20 (each s, 9H ꢅ 2, OPiv).

Anal. Calcd for C₃₂H₄₂O₁₀: C, 65.26; H, 7.22. Found: C, 65.26; H, 7.22.

p-[2-(Benzyloxycarbonyl)ethyl]phenyl 3,6-Di-O-pivaloyl-2,4-bis(O-tri-

fluoromethylsulfonyl)-ꢀ-D-galactopyranoside (11). To a solution of 10 (100

mg, 0.17 mmol) in pyridine (10 mL), trifluoromethanesulfonic anhydride (0.1 mL,

0.59 mmol) was added under argon at ꢃ19°C. The reaction mixture was stirred at

room temperature for 4 h until the disappearance of 10 on TLC with hexane/ethyl

acetate (2/1 v/v). The reaction mixture was poured into a saturated aq NaHCO₃so-

lution and extracted with CHCl₃. The combined organic layer was washed with

brine and water, dried over anhydrous magnesium sulfate, and concentrated to give

a residue. The remaining residue was purified on a short column of silica gel with

hexane/ethyl acetate (3/1 v/v) to afford 10 (145 mg, quantitatively): IR (cm^ꢃ1

)

1

2974 (OH), 1740 (CꢄO); H NMR (FX-200, CDCl₃) ꢈ 7.40–7.28 (m, 5H, -CH₂

Ph), 7.12, 6.94 (each d, 4H, phenylene, J ꢄ 8.8 Hz), 5.36 (d, 1H, H-4, J_3,4

ꢄ 2.7 Hz), 5.23 (dd, 1H, H-2, J_2,3ꢄ 9.3 Hz), 5.22 (dd, 1H, H-3), 5.20 (d, 1H, H-

1, J_1,2ꢄ 6.8 Hz), 5.11 (s, 2H, -CH₂Ph), 4.37 (dd, 1H, H-6, J_6,6ꢂꢄ 10.5 Hz), 4.19

(dd, 1H, H-5, J_5,6ꢂꢄ 6.1 Hz), 4.08 (dd, 1H, H-6, J_5,6ꢄ 6.1 Hz), 2.94 (t, 2H,

-CH₂CH₂CO-, J ꢄ 7.8 Hz), 2.65 (t, 2H, -CH₂CH₂CO-), 1.31, 1.21 (each s, 9H ꢅ

2, OPiv).

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AKAI ET AL.

p-[2-(Benzyloxycarbonyl)ethyl]phenyl 2,4-Di-O-acetyl-3,6-di-O-pival-

oyl-ꢀ-D-mannopyranoside (12). The reaction mixture of 11 (145 mg, 0.17

mmol), cesium acetate (98 mg, 0.51 mmol, 3.0 mol equiv) and 18-crown-6 ether

(135 mg, 0.51 mmol) in dry benzene (30 mL) was stirred overnight under ultra-

sonication for 12 h until the disappearance of 11 on TLC with hexane/ethyl acetate

(3/1 v/v). The reaction mixture was diluted with ethyl acetate, washed with brine

and water, dried over anhydrous magnesium sulfate, filtered, and concentrated to

give a residue. The remaining residue was purified on a column of silica gel with

hexane/ethyl acetate (3/1 v/v) to afford 12 (99 mg, 87% yield): [ꢁ]_D²⁵ꢃ36.1° (c

0.7, CHCl₃); IR (cm^ꢃ1) 1740 (CꢄO); ¹H NMR (FX-200, CDCl₃) ꢈ 7.33–7.26 (m,

5H, -CH₂Ph), 7.08, 6.92 (each d, 4H, phenylene, J ꢄ 8.7 Hz), 5.69 (dd, 1H, H-2,

J_2,3ꢄ 3.0 Hz), 5.34 (t, 1H, H-4, J_4,5ꢄ 9.7 Hz), 5.17 (d, 1H, H-1, J_1,2ꢄ 1.2 Hz),

5.10 (s, 2H, -CH₂Ph), 5.10 (dd, 1H, H-3, J_3,4ꢄ 9.7 Hz), 4.30 (dd, 1H, H-6ꢂ, J_6,6ꢂ

ꢄ 12.2 Hz), 4.17 (dd, 1H, H6, J_5,6ꢄ 6.7 Hz), 3.86 (ddd, 1H, H-5, J_5,6ꢂꢄ 2.4 Hz),

2.91 (t, 2H, -CH₂CH₂CO-, J ꢄ 7.8 Hz), 2.62 (t, 2H, -CH₂CH₂CO-), 2.20, 2.05

(each s, 3H ꢅ 2, OAc), 1.22, 1.15 (each s, 9H ꢅ 2, OPiv).

Anal. Calcd for C₃₆H₄₆O₁₂: C, 50.98; H, 5.76. Found: C, 64.03; H, 7.25.

p-[2-(Benzyloxycarbonyl)ethyl]phenyl 4-O-Acetyl-3,6-di-O-pivaloyl-2-

O-trifluoromethylsulfonyl-ꢀ-D-glucopyranoside (13). To a solution of 11

(1.27 g, 1.5 mmol) in dry toluene (55 mL), cesium acetate (344 mg, 1.79 mmol)

and 18-crown-6 ether (474 mg, 1.79 mmol, 2.0 mol equiv) were added and stirred

for 4 h at rt until the disappearance of 11 on TLC with hexane/ethyl acetate (3/1

v/v). The reaction mixture was diluted with ethyl acetate, washed with brine and

water, dried over anhydrous magnesium sulfate, filtered, and concentrated to give

a residue. The remaining residue was purified on a column of silica gel with hex-

ane/ethyl acetate (3/1 v/v) to afford 17 (1.04 g, 92% yield): [ꢁ]_D²⁵ꢃ14.6° (c 0.9,

CHCl₃); IR (cm^ꢃ1) 1743 (CꢄO); ¹H NMR (FX-200, CDCl₃) ꢈ 7.37–7.29 (m, 5H,

-CH₂Ph), 7.10, 6.70 (each d, 4H, phenylene, J ꢄ 8.5 Hz), 5.50 (dd, 1H, H-3, J_3,4

ꢄ 9.4 Hz), 5.14 (d, 1H, H-1, J_1,2ꢄ 7.9 Hz), 5.13 (dd, 1H, H-4, J_4,5ꢄ 9.4 Hz), 5.10

(s, 2H, -CH₂Ph), 4.97 (dd, 1H, H-2, J_2,3ꢄ 9.5 Hz), 4.23 (dd, 1H, H-6ꢂ, J_5,6ꢂꢄ 2.1

Hz), 4.12 (dd, 1H, H-6, J_6,6ꢂꢄ 12.2 Hz), 3.89 (ddd, 1H, H-5, J_5,6ꢄ 6.4 Hz), 2.93

(t, 2H, -CH₂CH₂CO-, J ꢄ 7.8 Hz), 2.65 (t, 2H, -CH₂CH₂CO-), 2.04 (s, 3H, OAc),

1.22, 1.21 (each s, 9H ꢅ 2, OPiv).

Anal. Calcd for C₃₅H₄₃F₃O₁₂S: C, 56.44; H, 5.82. Found: C, 56.35; H, 6.15.

p-[2-(Benzyloxycarbonyl)ethyl]phenyl 4-O-Acetyl-2-azido-2-deoxy-3,6-

di-O-pivaloyl-ꢀ-D-mannopyranoside (14). To a solution of 13 (839 mg, 1.10

mmol) in dry benzene (30 mL), tetrabutylammonium azide (627 mg, 2.20 mmol)

was added and stirred for 12 h in a water bath under ultrasonication until the dis-

appearance of 13 on TLC with hexane/ethyl acetate (3/1 v/v). The reaction mixture

was diluted with ethyl acetate, washed with brine and water, dried over anhydrous

magnesium sulfate, and concentrated to give a residue. The remaining residue was

purified on a column of silica gel with hexane/ethyl acetate (3/1 v/v) to afford 14

(691 mg, 96% yield): mp 61.5–64°C (ether-hexane); [ꢁ]_D²⁵ꢉ45.0° (c 0.9, CHCl₃);

ORDER

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SYNTHESIS OF NEW GLYCOPOLYMERS

133

IR (cm^ꢃ1) 2112 (N₃), 1732 (CꢄO); ¹H NMR (A-500, CDCl₃) ꢈ 7.38–7.29 (m, 5H,

-CH₂Ph), 7.02, 6.94 (each d, 4H, phenylene, J ꢄ 8.5 Hz), 5.31 (dd, 1H, H-4, J_4,5

ꢄ 9.7 Hz), 5.20 (d, 1H, H-1, J_1,2ꢄ 1.2 Hz), 5.10 (s, 2H, -CH₂Ph), 5.02 (dd, 1H,

H-3, J_3,4ꢄ 9.7 Hz), 4.27 (dd, 1H, H-2, J_2,3ꢄ 3.6 Hz), 4.26 (dd, 1H, H-6, J_6,6ꢂ

ꢄ

11.9 Hz), 4.13 (dd, 1H, H-6ꢂ, J_5,6ꢂꢄ 6.7 Hz), 3.77 (1H, ddd, H-5, J_5,6ꢄ 2.4 Hz),

2.92 (t, 2H, -CH₂CH₂CO-, J ꢄ 7.8 Hz), 2.64 (t, 2H, -CH₂CH₂CO-), 2.04 (s, 3H,

OAc), 1.23, 1.21 (each s, 9H ꢅ 2, OPiv).

Anal. Calcd for C₃₄H₄₃N₃O₁₀: C, 62.47; H, 6.63; N, 6.43. Found: C, 62.75;

H, 6.88; N, 6.34.

p-(Benzyloxycarbonylethyl)phenyl 2-Acetamido-4-O-acetyl-2-deoxy-

3,6-di-O-pivaloyl-ꢀ-D-mannopyranoside (15). Compound 14 (733 mg, 1.12

mmol) was hydrogenolyzed in the presence of a catalytic amount of 5% Pd-C/H₂

in benzene (25 mL) for 1 h. The reaction mixture was then filtered through a pad

of celite. The filtrate was treated with acetic anhydride (0.5 mL) at rt for 30 min,

and concentrated to give a residue. The remaining residue was purified on a col-

umn of silica gel with hexane/ethyl acetate (2/1 v/v) to afford 15 (400 mg, 53%

yield) and recovered 14 (311 mg, 42% yield): mp 116–117°C (dec.) (ethanol-hex-

25

1

ane); [ꢁ]_Dꢃ5.6° (c 0.7, CHCl₃); IR (cm^ꢃ1) 1737 and 1680 (CꢄO); H NMR

(EX-270, CDCl₃) ꢈ 7.37–7.30 (m, 5H, -CH₂Ph), 7.08, 6.91 (each d, 4H, phenylene,

J ꢄ 8.5 Hz), 5.82 (d, 1H, NHAc, J_NH,2ꢄ 9.5 Hz), 5.26 (d, 1H, H-1, J_1,2ꢄ 2.1 Hz),

5.15 (t, 1H, H-4, J_4,5ꢄ 8.5 Hz), 5.10 (s, 2H, -CH₂Ph), 5.07 (dd, 1H, H-3, J_3,4

ꢄ 8.5 Hz), 4.94 (ddd, 1H, H-2, J_2,3ꢄ 3.9 Hz), 4.25 (dd, 1H, H-6, J_6,6ꢂꢄ 12.2 Hz),

4.02 (dd, 1H, H-6ꢂ, J_5,6ꢂꢄ 6.7 Hz), 3.89 (ddd, 1H, H-5, J_5,6ꢄ 3.7 Hz), 2.91 (t, 2H,

-CH₂CH₂CO-, J ꢄ 7.3 Hz), 2.63 (t, 2H, -CH₂CH₂CO-), 2.07, 2.06 (each s, 6H,

OAc), 1.19, 1.19 (each s, 9H ꢅ 2, OPiv).

Anal. Calcd for C₃₆H₄₇NO₁₁: C, 64.56; H, 7.07; N, 2.09. Found: C, 64.72; H,

7.14; N, 1.97.

p-[2-(Benzyloxycarbonyl)ethyl]phenyl 4-O-Acetyl-2-deoxy-3,6-di-O-pi-

valoyl-ꢀ-D-arabino-hexopyranoside (16). To a solution of 13 (1.01 g, 1.64

mmol) in dry benzene (30 mL), tetrabutylammonium tetrahydroborate (680 mg,

2.70 mmol) was added, and kept for 29 h in a water bath under ultrasonication un-

til the disappearance of 13 on TLC with hexane/1ethyl acetate (3/1 v/v). The reac-

tion mixture was poured into a saturated aq NaHCO₃solution, extracted with ethyl

acetate, washed with brine and water, dried over anhydrous magnesium sulfate,

and concentrated to give a residue. The remaining residue was purified on a col-

umn of silica gel with hexane/ethyl acetate (3/1 v/v) to afford 16 (784 mg, 52%

25

yield): mp 70–71°C (ethanol-hexane); [ꢁ]_Dꢃ30.1° (c 1.0, CHCl₃); IR (cm^ꢃ1

)

1737 (CꢄO); ¹H NMR (A-500, CDCl₃) ꢈ 7.37–7.29 (m, 5H, -CH₂Ph), 7.09, 6.93

(each d, 4H, phenylene, J ꢄ 8.5 Hz), 5.16 (dd, 1H, H-1, J_1,2eqꢄ 2.0 Hz, J_1,2ax

ꢄ 9.4 Hz), 5.10 (s, 2H, -CH₂Ph), 5.06 (ddd, 1H, H-3, J_3,4ꢄ 9.5 Hz, J_2eq,3ꢄ 4.6

Hz), 5.04 (dd, 1H, H-4, J_4,5ꢄ 9.5 Hz), 4.24 (dd, 1H, H-6, J_5,6ꢄ 2.4 Hz, J_6,6ꢂ

ꢄ 12.1 Hz), 4.13 (dd, 1H, H-6ꢂ, J_5,6ꢂꢄ 6.9 Hz), 3.78 (ddd, 1H, H-5), 2.91 (t, 2H,

-CH₂CH₂CO-, J ꢄ 7.8 Hz), 2.64 (t, 2H, -CH₂CH₂CO-), 2.50 (ddd, 1H, H-2eq,

ORDER

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134

AKAI ET AL.

J

_2eq,2axꢄ 12.2 Hz), 2.05 (3H, s, OAc), 1.98 (1H, ddd, H-2ax), 1.21, 1.18 (each s,

9H ꢅ 2, OPiv).

Anal. Calcd for C₃₄H₄₄O₁₀: C, 66.65; H, 7.24. Found: C, 66.55; H, 7.24.

p-[2-(Benzyloxycarbonyl)ethyl]phenyl 4-O-Acetyl-2-deoxy-2-fluoro-3,6-

di-O-pivaloyl-ꢀ-D-mannopyranoside (17). To a solution of 13 (844 mg, 1.11

mmol) in dry benzene (50 mL), tetrabutylammonium fluoride (1M solution in

tetrahydrofuran, 5.6 mL, 0.55 mmol) was added dropwise at 0°C, and the reaction

mixture was stirred for 24 h at rt under argon until the disappearance of 13 on TLC

with hexane/ethyl acetate (3/1 v/v). The reaction mixture was poured into a satu-

rated aq NaHCO₃solution, extracted with ethyl acetate, washed with brine and wa-

ter, dried over anhydrous magnesium sulfate, filtered, and concentrated to give a

residue. The remaining residue was purified on a column of silica gel with hex-

ane/ethyl acetate (3/1 v/v) to afford 17 (258 mg, 37% yield): [ꢁ]_D²⁵ꢃ39.0° (c 0.8,

CHCl₃); IR (cm_ꢃ1) 1737 (CꢄO); ¹H NMR (A-500, CDCl₃) ꢈ 7.37–7.29 (m, 5H,

-CH₂Ph), 7.08, 6.91 (each d, 4H, styryl phenylene, J ꢄ 8.6 Hz), 5.41 (dd, 1H, H-4,

J

_3,4ꢄ J_4,5ꢄ 9.8 Hz), 5.12 (dd, 1H, H-1, J_1,2ꢄ 0.3 Hz, J_1,Fꢄ 15.2 Hz), 5.10 (s, 2H,

-CH₂Ph), 5.02 (ddd, 1H, H-3, J_2,3ꢄ 2.4 Hz, J_3,Fꢄ 23.8 Hz), 5.00 (ddd, 1H, H-2,

_2,Fꢄ 51.8 Hz), 4.30 (dd, 1H, H-6, J_5,6ꢄ 2.5 Hz, J_6,6ꢂꢄ 12.2 Hz), 4.16 (dd, 1H, H-

J

6, J_5,6ꢂꢄ 6.7 Hz), 3.83 (ddd, 1H, H-5), 2.92 (t, 2H, -CH₂CH₂CO-, J ꢄ 7.9 Hz), 2.64

(t, 2H, -CH₂CH₂CO-), 2.07 (s, 3H, OAc), 1.22, 1.21 (each s, 9H ꢅ 2, OPiv).

Anal. Calcd for C₃₄H₄₃FO₁₀; C, 64.75; H, 6.87. Found: C, 65.22; H, 7.01.

p-(2-Carboxyethyl)phenyl

2,4-Di-O-acetyl-3,6-di-O-pivaloyl-ꢀ-D-

mannopyranoside (18). p-[2-(Benzyloxycarbonyl)ethyl]phenyl glycoside 12

(754 mg, 1.12 mmol) was hydrogenolyzed in ethanol (25 mL) in the presence of a

catalytic amount of 10% Pd(OH)₂-C/H₂for 2 h until the disappearance of 12 on

TLC with hexane/ethyl acetate (3/1 v/v). The reaction mixture was filtered through

a pad of celite and concentrated to afford 18 (617 mg, 95% yield): [ꢁ]_D²⁵ꢃ44.4°

(c 0.8, CHCl₃); IR (cm^ꢃ1) 3240 (OH), 1746 (CꢄO); ¹H NMR (FX-200, CDCl₃) ꢈ

7.07, 6.92 (each d, 4H, phenylene, J ꢄ 7.6 Hz), 5.67 (dd, 1H, H-2, J_2,3ꢄ 3.2 Hz),

5.34 (t, 1H, H-4, J_4,5ꢄ 9.8 Hz), 5.19 (s, 1H, H-1), 5.10 (dd, 1H, H-3, J_3,4ꢄ 9.8

Hz), 4.30 (dd, 1H, H-6ꢂ, J_6,6ꢂꢄ 12.0 Hz), 4.19 (dd, 1H, H-6), 3.84 (ddd, 1H, H-5,

J_5,6ꢄ 6.6 Hz), 2.85 (t, 2H, -CH₂CH₂CO-, J ꢄ 7.8 Hz), 2.58 (t, 2H, -CH₂CH₂

CO-), 2.20, 2.05 (each s, 6H, OAc), 1.22, 1.15 (each s, 9H ꢅ 2, OPiv).

Anal. Calcd for C₂₈H₄₀O₁₂: C, 59.14; H, 7.09. Found: C, 59.64; H, 7.08.

p-(2-Carboxyethyl)phenyl 2-Acetamido-4-O-acetyl-2-deoxy-3,6-di-O-pi-

valoyl-ꢀ-D-mannopyranoside (19). p-[2-(Benzyloxycarbonyl)ethyl]phenyl

glycoside 15 (80 mg, 0.12 mmol) was hydrogenolyzed in ethanol (10 mL) in the

presence of a catalytic amount of 10% Pd(OH)₂-C/H₂for 1 h until the disappear-

ance of 15 on TLC with hexane/ethyl acetate (2/1 v/v). The reaction mixture was

then filtered through a pad of celite and concentrated to afford 19 (63 mg, 91%

yield): mp 88–91°C (ethanol-hexane); [ꢁ]_Dꢃ55.6° (c 0.6, CHCl₃); IR (cm^ꢃ1

25

)

3388 (OH), 1728 and 1644 (CꢄO); ¹H NMR (A-500, CDCl₃) ꢈ 7.10, 6.93 (each d,

ORDER

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SYNTHESIS OF NEW GLYCOPOLYMERS

135

4H, phenylene, J ꢄ 8.5 Hz), 6.80 (br s, 1H, COOH), 5.89 (d, 1H, NHAc, J_NH,2

ꢄ 9.1 Hz), 5.27 (d, 1H, H-1, J_1,2ꢄ 1.9 Hz), 5.15 (t, 1H, H-4, J_4,5ꢄ 8.6 Hz), 5.08

(dd, 1H, H-3, J_3,4ꢄ 8.6 Hz), 4.95 (ddd, 1H, H-2, J_2,3ꢄ 4.0 Hz), 4.24 (dd, 1H, H-

6, J_6,6ꢂꢄ 12.2 Hz), 4.20 (dd, 1H, H-6ꢂ, J_6ꢂ,5ꢄ 6.4 Hz), 3.90 (ddd, 1H, H-5, J_5,6

ꢄ 3.6 Hz), 2.89 (t, 2H, -CH₂CH₂CO-, J ꢄ 7.7 Hz), 2.62 (t, 2H, -CH₂CH₂CO-),

2.07, 2.06 (each s, 3H ꢅ 2, OAc), 1.19, 1.19 (each s, 9H ꢅ 2, OPiv).

Anal. Calcd for C₂₉H₄₁NO₁₁: C, 60.09; H, 7.13; N, 2.42. Found: C, 59.80; H,

6.64; N, 2.47.

p-(2-Carboxyethyl)phenyl 4-O-Acetyl-2-deoxy-3,6-di-O-pivaloyl-ꢀ-D-

arabino-hexopyranoside (20). p-[2-(Benzyloxycarbonyl)ethyl]phenyl glyco-

side 16 (610 mg, 1.00 mmol) was hydrogenolyzed in ethanol (50 mL) in the

presence of a catalytic amount of 10% Pd(OH)₂-C/H₂for 1 h until the disappear-

ance of 16 on TLC with hexane/ethyl acetate (3/1 v/v). The reaction mixture was

then filtered through a pad of celite and concentrated to afford 20 (476 mg, 92%

yield): mp 147–148°C (ethanol-hexane); [ꢁ]_D²⁵ꢃ35.0° (c 1.0, CHCl₃); IR (cm^ꢃ1

)

3460 (OH), 1734 (CꢄO); ¹H NMR (A-500, CDCl₃) ꢈ 8.58 (br s, 1H, COOH), 7.09,

6.95 (each d, 4H, -CH₂Ph, J ꢄ 8.9 Hz), 5.17 (dd, 1H, H-1, J_1,2eqꢄ 2.2 Hz, J_1,2ax

ꢄ 9.3 Hz), 5.06 (ddd, 1H, H-3, J_3,4ꢄ 9.4 Hz, J_2eq,3ꢄ 4.6 Hz, J_2ax,3ꢄ 11.2 Hz),

5.04 (dd, 1H, H-4, J_4,5ꢄ 9.5 Hz), 4.24 (dd, 1H, H-6, J_5,6ꢄ 2.4 Hz, J_6,6ꢂꢄ 12.1

Hz), 4.13 (dd, 1H, H-6ꢂ, J_5,6ꢂꢄ 6.9 Hz), 3.79 (ddd, 1H, H-5), 2.89 (t, 2H,

-CH₂CH₂CO-, J ꢄ 7.8 Hz), 2.62 (t, 2H, -CH₂CH₂CO-), 2.50 (ddd, 1H, H-2eq,

J

_2eq,2axꢄ 12.2 Hz), 2.05 (s, 3H, OAc), 1.98 (ddd, 1H, H-2ax), 1.21, 1.18 (each s,

9H ꢅ 2, OPiv).

Anal. Calcd for C₂₇H₃₈O₁₀: C, 62.05; H, 7.33. Found: C, 62.26; H, 7.33.

p-(2-Carboxyethyl)phenyl 4-O-Acetyl-2-deoxy-2-fluoro-3,6-di-O-pival-

oyl-ꢀ-D-mannopyranoside (21). p-[2-(Benzyloxycarbonyl)ethyl]phenyl glyco-

side 17 (224 mg, 0.36 mmol) was hydrogenolyzed in ethanol (50 mL) in the

presence of a catalytic amount of 10% Pd(OH)₂-C/H₂for 1 h until the disappear-

ance of 17 on TLC with hexane/ethyl acetate (2/1 v/v). The reaction mixture was

then filtered through a pad of celite and concentrated to afford 21 (173 mg, 90%):

mp 168–169°C (ethanol-hexane); [ꢁ]_D²⁵ꢃ40.3° (c 1.0, CHCl₃); IR (cm^ꢃ1) 3358

(OH), 1740 (CꢄO); ¹H NMR (A-500, CDCl₃) ꢈ 9.25 (br s, 1H, COOH), 7.12, 6.99

(each d, 4H, phenylene, J ꢄ 8.5 Hz), 5.41 (dd, 1H, H-4, J_3,4ꢄ J_4,5ꢄ 9.8 Hz), 5.15

(dd, 1H, H-1, J_1,2ꢄ 0.3 Hz, J_1,Fꢄ 17.1 Hz), 5.03 (ddd, 1H, H-3, J_2,3ꢄ 2.5 Hz, J_3,F

ꢄ 26.8 Hz), 5.01 (ddd, 1H, H-2, J_2,Fꢄ 51.9 Hz), 4.30 (dd, 1H, H-6, J_5,6ꢄ 2.4 Hz,

J_6,6ꢂꢄ 12.2 Hz), 4.17 (dd, 1H, H-6ꢂ, J_5,6ꢂꢄ 6.7 Hz), 3.85 (ddd, 1H, H-5), 2.90 (t,

2H, -CH₂CH₂CO-, J ꢄ 7.7 Hz), 2.63 (t, 2H, -CH₂CH₂CO-), 2.06 (s, 3H, OAc),

1.22 (each s, 9H ꢅ 2, OPiv).

Anal. Calcd for C₂₇H₃₇FO₁₀: C, 60.00; H, 6.90. Found: C, 60.07; H, 7.13.

p-[2-(N-p-Vinylbenzyl)carbamoyl]ethyl]phenyl 2,4-Di-O-acetyl-3,6-di-

O-pivaloyl-ꢀ-D-mannopyranoside (22). To a solution of p-(2-carboxyethyl)

phenyl glycoside 18 (617 mg, 1.06 mmol) and 1-ethyl-3-(3-dimethylamino-

ORDER

REPRINTS

136

AKAI ET AL.

propyl)carbodiimide (WSC) (275 mg, 1.43 mmol) in dichloromethane (50 mL), a

solution of p-vinylbenzylamine¹⁶(200 mg, 1.52 mmol) in dichloromethane (20

mL) was added dropwise at 0°C, and the reaction mixture was stirred for 1 h until

the disappearance of 18 on TLC with hexane/ethyl acetate (3/1 v/v). The reaction

mixture was diluted with dichloromethane, washed with 10% aq citric acid solu-

tion, brine and water, dried over anhydrous magnesium sulfate, filtered, and

concentrated to give a residue. The remaining residue was purified on a column of

silica gel with hexane/ethyl acetate (3/5 v/v) to afford 22 (496 mg, 67% yield): mp

25

125–129°C (ethanol-hexane); [ꢁ]_Dꢃ33.0° (c 0.8, CHCl₃); IR (cm^ꢃ1) 1736

(CꢄO), 1648 (CꢄC); ¹H NMR (FX-200, CDCl₃) ꢈ 7.34, 7.07 (each d, 4H, styryl

phenylene, J ꢄ 8.3 Hz), 7.11, 6.92 (each d, 4H, phenylene, J ꢄ 8.5 Hz), 6.69 (dd,

1H, -Ph-CHꢄCHaHb, J_transꢄ 17.6 Hz, J_cisꢄ 11.0 Hz), 5.74 (dd, 1H, Ha, J_gem

ꢄ 0.9 Hz), 5.68 (dd, 1H, H-2, J_2,3ꢄ 3.2 Hz), 5.60 (t, 1H, -CONH-), 5.34 (t, 1H,

H-4, J_4,5ꢄ 9.8 Hz), 5.25 (dd, 1H, Hb), 5.17 (d, 1H, H-1, J_1,2ꢄ 1.0 Hz), 5.10 (dd,

1H, H-3, J_3,4ꢄ 9.8 Hz), 4.36 (d, 2H, -NHCH₂-, J_NH,CH2ꢄ 5.6 Hz), 4.30 (dd, 1H,

H-6ꢂ, J_5,6ꢂꢄ 7.1 Hz, J_6,6ꢂꢄ 12.0 Hz), 4.16 (dd, 1H, H-6), 3.84 (ddd, 1H, H-5, J_5,6

ꢄ 2.4 Hz), 2.93 (t, 2H, -CH₂CH₂CO-, J ꢄ 7.8 Hz), 2.46 (t, 2H, -CH₂CH₂CO-),

2.20, 2.05 (each s, 3H ꢅ 2, OAc), 1.21, 1.15 (each s, 9H ꢅ 2, OPiv).

Anal. Calcd for C₃₈H₄₉O₁₁N: C, 59.14; H, 7.09, N, 2.39. Found: C, 65.82; H,

7.26, N, 2.01.

p-[2-[N-(p-Vinylbenzyl)carbamoyl]ethyl]phenyl 2-Acetamido-4-O-

acetyl-2-deoxy-3,6-di-O-pivaloyl-ꢀ-D-mannopyranoside (23). To a solution

of p-(2-carboxyethyl)phenyl glycoside 19 (170 mg, 0.29 mmol) and 1-ethyl-3-(3-

dimethylaminopropyl)carbodiimide (WSC) (67 mg, 0.35 mmol) in dichlo-

romethane (25 mL), a solution of p-vinylbenzylamine¹⁶(5 mg, 0.38 mmol) in

dichloromethane (10 mL) was added dropwise at 0°C, and the reaction mixture was

stirred for 1 h until the disappearance of 19 on TLC with hexane/ethyl acetate (1/3

v/v). The reaction mixture was diluted with dichloromethane, washed with 10% aq

citric acid solution, brine and water, dried over anhydrous magnesium sulfate, fil-

tered, and concentrated to give a residue. The remaining residue was purified on a

column of silica gel with hexane/ethyl acetate/pyridine (25/75/3 v/v/v) to afford 23

25

(124 mg, 81% yield): mp 88–91°C (hexane-ethanol): [ꢁ]_Dꢃ44.3° (c 0.9,

CHCl₃); IR (cm^ꢃ1) 1737 (CꢄO), 1653 (CꢄC); ¹H NMR (A-500, CDCl₃) ꢈ 7.34,

7.11 (each d, 4H, styryl phenylene, J ꢄ 8.3 Hz), 7.09, 6.90 (each d, 4H, phenylene,

J ꢄ 8.5 Hz), 6.70 (dd, 1H, -Ph-CHꢄCHaHb, J_transꢄ 17.4 Hz, J_cisꢄ 10.9 Hz), 5.84

(d, 1H, NHAc, J_NH,2ꢄ 9.2 Hz), 5.73 (dd, 1H, Ha, J_gemꢄ 0.9 Hz), 5.65 (t, 1H, -

CONH-), 5.25 (dd, 1H, Hb), 5.24 (d, 1H, H-1, J_1,2ꢄ 1.8 Hz), 5.15 (t, 1H, H-4, J_4,5

ꢄ 8.8 Hz), 5.07 (dd, 1H, H-3, J_3,4ꢄ 8.9 Hz), 4.94 (ddd, 1H, H-2, J_2,3ꢄ 3.9 Hz),

4.38 (1H, d, -NHCH₂-, J_NH,CH2ꢄ 5.6 Hz), 4.25 (dd, 1H, H-6, J_6,6ꢂꢄ 11.9 Hz), 4.19

(dd, 1H, H-6ꢂ, J_5,6ꢂꢄ 6.7 Hz), 3.88 (ddd, 1H, H-5, J_5,6ꢄ 3.4 Hz), 2.94 (t, 2H,

-CH₂CH₂CO-, J ꢄ 7.9 Hz), 2.64 (t, 2H, -CH₂CH₂CO-), 2.07, 2.06 (each s, 3H ꢅ

2, OAc), 1.19, 1.19 (each s, 9H ꢅ 2, OPiv); ¹³C NMR (A-500, CDCl₃) ꢈ 177.91,

177.29, 171.71, 170.21, 169.51 (CꢄO), 135.66 (CHꢄCH₂), 129.51, 128.34,

ORDER

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SYNTHESIS OF NEW GLYCOPOLYMERS

137

128.04, 126.50, 116.80 (Ph), 114.09 (CHꢄCH₂), 97.00 (C-1), 72.75 (C-5), 70.77

(C-3), 66.18 (C-4), 62.72 (C-6), 49.66 (C-2), 43.34, 38.81, 30.88 (CH₂).

Anal. Calcd for C₃₈H₅₀N₂O₁₀: C, 65.68; H, 7.24; N, 4.03. Found: C, 65.18;

H, 7.36; N, 3.86.

p-[2-[N-(p-Vinylbenzyl)carbamoyl]ethyl]phenyl 4-O-Acetyl-2-deoxy-

3,6-di-O-pivaloyl-ꢀ-D-arabino-hexopyranoside (24). To a solution of p-(car-

boxyethyl)phenyl glycoside 20 (60 mg, 0.11 mmol) and 1-ethyl-3-(3-dimethy-

laminopropyl)carbodiimide (WSC) (24 mg, 0.12 mmol) in dichloromethane (10

mL), a solution of p-vinylbenzylamine¹⁶(5 mg, 0.15 mmol) in dichloromethane (3

mL) was added dropwise at 0°C, and the reaction mixture was stirred for 1 h until

the disappearance of 20 on TLC with hexane/ethyl acetate (1/2 v/v). The reaction

mixture was diluted with dichloromethane, washed with 10% aq citric acid

solution, brine and water, dried over anhydrous magnesium sulfate, filtered, and

concentrated to give a residue. The remaining residue was purified on a column of

silica gel with hexane/ethyl acetate (1/1 v/v) to afford 24 (56 mg, 76% yield): mp

25

131–132°C (ethanol-hexane); [ꢁ]_Dꢃ24.4° (c 1.0, CHCl₃); IR (cm^ꢃ1) 1734

1

(CꢄO), 1644 (CꢄC); H NMR (A-500, CDCl₃) ꢈ 7.34, 7.11 (each d, 4H, styryl

phenylene, J ꢄ 8.0 Hz), 7.08, 6.92 (each d, 4H, phenylene, J ꢄ 8.6 Hz), 6.69 (dd,

1H, -Ph-CHꢄCHaHb, J_transꢄ 17.4 Hz, J_cisꢄ 10.7 Hz) 5.73 (dd, 1H, Ha, J_gemꢄ

0.6 Hz), 5.66 (t, 1H, -CONH-), 5.25 (dd, 1H, Hb), 5.14 (dd, 1H, H-1, J_1,2eqꢄ 2.1

Hz, J_1,2axꢄ 9.4 Hz), 5.05 (ddd, 1H, H-3, J_2eq,3ꢄ 4.9 Hz, J_2ax,3ꢄ 11.6 Hz, J_3,4ꢄ

9.1 Hz), 5.04 (dd, 1H, H-4, J_4,5ꢄ 9.1 Hz), 4.37 (d, 2H, -NHCH₂-, J_NH,CH2ꢄ 5.6

Hz), 4.24 (dd, 1H, H-6, J_5,6ꢄ 2.2 Hz, J_6,6ꢂꢄ 11.9 Hz), 4.12 (dd, 1H, H-6ꢂ, J_5,6ꢂ

ꢄ

6.7 Hz), 3.78 (ddd, 1H, H-5), 2.93 (t, 2H, -CH₂CH₂CO-, J ꢄ 7.8 Hz), 2.50 (ddd,

1H, H-2, J_2eq,2axꢄ 11.3 Hz), 2.46 (t, 2H, -CH₂CH₂CO-), 2.05 (s, 3H, OAc), 1.98

(ddd, 1H, H-2ax), 1.20, 1.18 (each s, 9H ꢅ 2, OPiv); ¹³C NMR (A-500, CDCl₃) ꢈ

178.26, 177.69, 171.78, 169.53 (CꢄO), 136.36 (CHꢄCH₂), 129.36, 116.83,

127.99, 126.50 (Ph), 114.06 (CHꢄCH₂), 97.42 (C-1), 72.49 (C-5), 70.12 (C-3),

68.92 (C-4), 63.01 (C-6), 35.90 (C-2), 43.32, 38.73, 30.93 (CH₂).

Anal. Calcd for C₃₆H₄₇O₉N: C, 67.80; H, 7.43; N, 2.20. Found: C, 68.24; H,

7.72; N, 2.00.

p-[2-[N-(p-Vinylbenzyl)carbamoyl]ethyl]phenyl 4-O-Acetyl-2-deoxy-2

-fluoro-3,6-di-O-pivaloyl-ꢀ-D-mannopyranoside (25). To a solution of p-(pro-

pionic acid)phenyl glycoside 21 (83 mg, 0.15 mmol) and 1-ethyl-3-(3-dimethy-

laminopropyl)carbodiimide (WSC) (32 mg, 0.16 mmol) in dichloromethane (5

mL), a solution of p-vinylbenzylamine¹⁶(25 mg, 0.19 mmol) in dichloromethane

(5 mL) was added dropwise at 0°C, and the reaction mixture was stirred for 20 min

until the disappearance of 21 on TLC with hexane/ethyl acetate (1/3 v/v). The re-

action mixture was diluted with dichloromethane, washed with 10% aq citric acid

solution, brine and water, dried over anhydrous magnesium sulfate, filtered, and

concentrated to give a residue. The remaining residue was purified on a column of

silica gel with hexane/ethyl acetate (1/2 v/v) to afford 25 (62 mg, 61%): mp

ORDER

REPRINTS

138

AKAI ET AL.

25

86–89°C (ether-hexane); [ꢁ]_Dꢃ35.2° (c 0.9, CHCl₃); IR (cm^ꢃ1) 1737 (CꢄO)

and 1646 (CꢄC); ¹H NMR (A-500, CDCl₃) ꢈ 7.34, 7.11 (each d, 4H, styryl pheny-

lene, J ꢄ 7.9 Hz), 7.09, 6.96 (each d, 4H, phenylene, J ꢄ 9.2 Hz), 6.69 (dd, 1H,

-Ph-CHꢄCHaHb, J_transꢄ 17.7 Hz, J_cisꢄ 11.0 Hz), 5.74 (dd, 1H, Ha, J_gemꢄ 0.6

Hz), 5.61 (t, 1H, -CONH-), 5.41 (d, 1H, H-4, J_3,4ꢄ J_4,5ꢄ 9.8 Hz), 5.25 (dd, 1H,

Hb), 5.10 (dd, 1H, H-1, J_1,2ꢄ 0.3 Hz, J_1,Fꢄ 17.1 Hz), 5.02 (dd, 1H, H-3, J_2,3

ꢄ 2.4 Hz, J_3,Fꢄ 26.9 Hz), 5.00 (ddd, 1H, H-2, J_2,Fꢄ 52.3 Hz), 4.37 (d, 1H, -

NHCH₂-, J_NH,CH2ꢄ 5.6 Hz), 4.29 (dd, 1H, H-6, J_5,6ꢄ 2.3 Hz, J_6,6ꢂꢄ 11.9 Hz),

4.15 (dd, 1H, H-6ꢂ, J_{5, 6ꢂ}ꢄ 6.8 Hz), 3.83 (ddd, 1H, H-5), 2.94 (t, 2H, -CH₂CH₂

CO-, J ꢄ 7.4 Hz), 2.46 (t, 2H, -CH₂CH₂CO-), 2.06 (s, 3H, OAc), 1.22, 1.20 (each

s, 9H ꢅ 2, OPiv); ¹³C NMR (A-500, CDCl₃) ꢈ 178.17, 177.61, 171.69, 169.54

(CꢄO), 136.33 (CHꢄCH₂), 129.47, 127.97, 126.50, 116.82 (Ph), 114.09

(CHꢄCH₂), 96.84 (C-1, J_1,Fꢄ 15.5 Hz), 87.01 (C-2, J_2,Fꢄ 191.4 Hz), 72.77 (C-

5), 71.40 (C-3, J_3,Fꢄ 16.6 Hz), 65.63 (C-4), 62.56 (C-6), 43.30, 38.65, 30.90

(CH₂).

Anal. Calcd for C₃₇H₄₅FO₉N₁: C, 66.65; H, 6.80; N, 2.10. Found: C, 66.50;

H, 6.70; N, 1.95.

p-[2-[N-(p-Vinylbenzyl)carbamoyl]ethyl]phenyl ꢀ-D-Mannopyranoside

(26). To a solution of p-[2-[N-(p-vinylbenzyl)carbamoyl]ethyl]phenyl glycoside

22 (484 mg, 0.69 mmol) in methanol (10 mL), sodium methoxide (1.9M in

methanol solution) was carefully added at pH 10, and the reaction mixture was

stirred for 2 h until the disappearance of 22 on TLC with hexane/ethyl acetate (1/2

v/v). The reaction mixture was neutralized with Dowex 50W-X8 (H^ꢉform) ion-

exchange resin, filtered, and concentrated to afford 26 (289 mg, 94% yield): mp

123–126°C (methanol); [ꢁ]_D²⁵ꢃ26.8° (c 0.7, MeOH); IR (cm^ꢃ1) 3322 (OH), 1644

(CꢄC); ¹H NMR (A-500, d₆-DMSO) ꢈ 8.23 (t, 1H, NH), 7.36, 7.12 (each d, 4H,

styryl phenylene, Hb, J_transꢄ 17.7 Hz, J_cisꢄ 11.0 Hz), 5.75 (dd, 1H, Ha, J_gem

ꢄ 0.9 Hz), 5.22 (dd, 1H, Hb), 5.22 (s, 1H, H-1), 4.24 (d, 2H, -NHCH₂-, J_NH,CH2

ꢄ 5.1 Hz), 3.88 (d, 1H, H-2, J_2,3ꢄ 3.3 Hz), 3.73 (dd, 1H, H-3, J_3,4ꢄ 11.3 Hz),

3.51–3.41 (m, 3H, H-4, H-6, and H-6ꢂ), 3.32 (br s, 4H, OH), 3.24 (ddd, 1H, H-5,

J

_4,5ꢄ 8.8 Hz, J_5,6ꢄ 2.1 Hz, J_5,6ꢂꢄ 6.1 Hz), 2.81 (t, 2H, -CH₂CH₂CO-, J ꢄ 7.8

Hz), 2.42 (t, 2H, -CH₂CH₂CO-); ¹³C NMR (A-500, d₆-DMSO) ꢈ 171.24 (CꢄO),

136.46, 135.52, 134.20, 128.94, 127.25, 125.79 (Ph), 136.22 (CHꢄCH₂), 113.46

(CHꢄCH₂), 97.84 (C-1), 77.43 (C-2), 73.42 (C-3), 70.54 (C-4), 66.78 (C-5), 60.99

(C-6), 41.70, 37.16, 30.22 (CH₂).

Anal. Calcd for C₂₄H₂₉O₇N: C, 65.00; H, 6.59; N, 3.16. Found: C, 64.94; H,

6.48; N, 2.99.

p-[2-[N-(p-Vinylbenzyl)carbamoyl]ethyl]phenyl 2-Acetamido-2-deoxy-

ꢀ-D-mannopyranoside (27). To a solution of p-[2-[N-(p-vinylbenzyl)car-

bamoyl]ethyl]phenyl glycoside 23 (478 mg, 0.69 mmol) in methanol (30 mL),

sodium methoxide (1.9M in methanol solution) was carefully added at pH 10, and

the reaction mixture was stirred for 2 h until the disappearance of 23 on TLC with

hexane/ethyl acetate (1/4 v/v). The reaction mixture was neutralized with Dowex

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139

50W-X8 (H^ꢉform) ion-exchange resin, filtered, and concentrated to afford 27

(333 mg, quantitatively): mp 99–102°C (methanol); [ꢁ]_D²⁵ꢃ67.8° (c 0.8, MeOH);

IR (cm^ꢃ1) 3304 (OH), 1656 (CꢄC); ¹H NMR (A-500, d₆-DMSO) ꢈ 8.29 (t, 1H, -

CONH-), 7.42 (d, 1H, NHAc, J_NH,2ꢄ 9.8 Hz), 7.37, 7.12 (each d, 4H, styryl

phenylene, J ꢄ 8.3 Hz), 7.09, 6.86 (each d, 4H, phenylene, J ꢄ 8.5 Hz), 6.70 (dd,

1H, -Ph-CHꢄCHaHb, J_transꢄ 17.4 Hz, J_cisꢄ 10.9 Hz), 5.78 (dd, 1H, Ha, J_gem

ꢄ 0.6 Hz ), 5.21 (dd, 1H, Hb,), 5.19 (d, 1H, H-1, J_1,2ꢄ 1.5 Hz), 4.47 (ddd, 1H, H-

2, J_2,3ꢄ 1.8 Hz), 4.22 (d, 2H, -NHCH₂-, J_NH,CH2ꢄ 5.1 Hz), 3.74 (dd, 1H, H-3, J_3,4

ꢄ 11.6 Hz), 3.57 (dd, 1H, H-6, J_6,6ꢂꢄ 9.2 Hz), 3.54 (dd, 1H, H-6ꢂ, J_6ꢂ,5ꢄ 6.4 Hz),

3.34 (t, 1H, H-4, J_4,5ꢄ 9.7 Hz), 3.29 (ddd, 1H, H-5, J_5,6ꢄ 4.6 Hz), 3.29 (br s, 4H,

OH), 2.77 (t, 2H, -CH₂CH₂CO-, J ꢄ 7.9 Hz), 2.40 (t, 2H, -CH₂CH₂CO-), 1.93 (s,

3H, NHAc); ¹³C NMR (A-500, d₆-DMSO) ꢈ 171.27, 169.97 (CꢄO), 129.07,

127.37, 125.95, 116.06 (Ph), 134.51 (CHꢄCH₂), 113.75 (CHꢄCH₂), 96.94 (C-1),

77.87 (C-5), 73.97 (C-3), 66.97 (C-4), 61.04 (C-6), 52.35 (C-2), 41.70, 37.11,

30.22 (CH₂), 22.94 (CH₃).

Anal. Calcd for C₂₆H₃₂N₂O₇: C, 64.45; H, 6.66; N, 5.78. Found: C, 64.34; H,

6.53; N, 5.84.

p-[2-[N-(p-Vinylbenzyl)carbamoyl]ethyl]phenyl ꢀ-D-Arabino-hexopyra-

noside (28). To a solution of p-[2-[N-(p-vinylbenzyl)carbamoyl]ethyl]phenyl

glycoside 24 (416 mg, 0.65 mmol) in methanol (18 mL), sodium methoxide (1.9M

in methanol solution) was carefully added at pH 10, and the reaction mixture was

stirred for 2 h until the disappearance of 24 on TLC with hexane/ethyl acetate (1/1

v/v). The reaction mixture was neutralized with Dowex 50W-X8 (H^ꢉform) ion-

exchange resin, filtered, and concentrated to afford 28 (279 mg, quantitatively): mp

171–172°C (ethanol-hexane); [ꢁ]_Dꢃ28.6° (c 0.6, DMSO); IR (cm^ꢃ1) 3298

25

1

(OH), 1644 (CꢄC); H NMR (A-500, d₆-DMSO) ꢈ 8.28 (t, 1H, -CONH-), 7.37,

7.09 (each d, 4H, styryl phenylene, J ꢄ 8.3 Hz), 7.11, 6.90 (each d, 4H, phenylene,

J ꢄ 8.5 Hz), 6.69 (dd, 1H, -Ph-CHꢄCHaHb, J_transꢄ 17.7 Hz, J_cisꢄ 11.0 Hz), 5.78

(dd, 1H, Ha, J_gemꢄ 0.9 Hz), 5.21 (dd, 1H, Hb), 5.16 (dd, 1H, H-1, J_1,2eqꢄ 2.2 Hz,

J

_1,2axꢄ 9.7 Hz), 4.22 (d, 2H, -NHCH₂-, J_NH,CH2ꢄ 5.8 Hz), 3.68 (ddd, 1H, H-6,

_5,6ꢄ 2.1 Hz, J_6,6ꢂꢄ 11.9 Hz), 3.50 (ddd, 1H, H-6ꢂ, J_5,6ꢄ 6.0 Hz), 3.48 (dd, 1H,

H-4, J_5,4ꢄ J_5,6ꢄ 9.5 Hz), 3.54 (dd, 1H, H-6 ꢂ, J_5,6ꢂꢄ 6.4 Hz), 3.34 (t, 1H, H-4,

_4,5ꢄ 9.7 Hz), 3.31 (br s, 3H, OH), 3.23 (ddd, 1H, H-5), 3.05 (ddd, 1H, H-3, J_2eq,3

J

ꢄ 5.2 Hz, J_2ax,3ꢄ 11.6 Hz), 2.77 (t, 2H, -CH₂CH₂CO-, J ꢄ 7.8 Hz), 2.40 (t, 2H,

-CH₂CH₂CO-), 2.14 (ddd, 1H, H-2ax, J_2ax,2eqꢄ 12.3 Hz), 1.55 (ddd, 1H, H-2eq);

¹³C NMR (A-500, d₆-DMSO) ꢈ 171.25 (CꢄO), 136.35 (CHꢄCH₂), 129.05,

127.34, 125.93, 115.87 (Ph), 113.72 (CHꢄCH₂), 96.73 (C-1), 77.18 (C-5), 71.21

(C-3), 70.38 (C-4), 60.82 (C-6), 41.67 (NHCH₂), 39.50 (C-2), 37.15, 30.2 (CH₂).

Anal. Calcd for C₂₄H₂₉O₆N: C, 67.80; H, 6.84; N, 3.28. Found: C, 67.17; H,

7.03; N, 3.31.

p-[2-[N-(p-Vinylbenzyl)carbamoyl]ethyl]phenyl 2-Deoxy-2-fluoro-ꢀ-D-

mannopyranoside (29). To a solution of p-[2-[N-(p-vinylbenzyl)carbamoyl]-

ethyl]phenyl glycoside 25 (37 mg, 0.06 mmol) in methanol (1 mL), sodium

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AKAI ET AL.

methoxide (1.9M in methanol solution, 1 mL) was carefully added at pH 10, and

the reaction mixture was stirred for 2 h until the disappearance of 25 on TLC with

hexane/ethyl acetate (1/2 v/v). The reaction mixture was neutralized with Dowex

50W-X8 (H^ꢉform) ion-exchange resin, filtered, and concentrated to afford 29 (25

mg, quantitatively): mp 185–186°C (ether-hexane); [ꢁ]_D²⁵ꢃ35.2° (c 0.2, DMSO);

IR (cm^ꢃ1) 3280 (OH), 1641 (CꢄC); ¹H NMR (A-500, d₆-DMSO) ꢈ 8.30 (t, 1H, -

CONH-), 7.38, 7.12 (each d, 4H, styryl phenylene, J ꢄ 7.9 Hz), 7.12, 6.92 (each d,

4H, phenylene, J ꢄ 8.6 Hz), 6.69 (dd, 1H, -Ph-CHꢄCHaHb, J_transꢄ 17.8 Hz, J_cis

ꢄ 11.0 Hz), 5.78 (dd, 1H, Ha, J_gemꢄ 0.6 Hz), 5.28 (dd, 1H, H-1, J_1,2ꢄ 0.3 Hz,

J_1,Fꢄ 19.5 Hz), 5.21 (dd, 1H, Hb), 4.72 (ddd, 1H, H-2, J_2,Fꢄ 52.5 Hz), 4.22 (d,

2H, -NHCH₂-, J_NH,CH2ꢄ 4.9 Hz), 3.70 (dd, 1H, H-6, J_6,6ꢂꢄ 11.7 Hz), 3.57 (ddd,

1H, H-3, J_2,3ꢄ 2.4 Hz, J_3,Fꢄ 31.1 Hz), 3.47 (dd, 1H, H-6ꢂ, J_6ꢂ,5ꢄ 5.7 Hz), 3.40

(dd, 1H, H-4, J_3,4ꢄ J_4,5ꢄ 9.3 Hz), 3.29 (ddd, 1H, H-5, J_5,6ꢄ 1.2 Hz), 3.29 (br s,

3H, OH), 2.78 (t, 2H, -CH₂CH₂CO-, J ꢄ 7.8 Hz), 2.40 (t, 1H, -CH₂CH₂CO-); ¹³

C

NMR (A-500, d₆-DMSO) ꢈ 171.24 (CꢄO), 136.34 (CHꢄCH₂), 129.13, 127.35,

125.95, 115.70 (Ph), 113.73 (CHꢄCH₂), 95.75 (C-1, J_1,Fꢄ 14.4 Hz), 90.53 (C-2,

J_2,Fꢄ 185.2 Hz), 77.47 (C-5), 71.70 (C-3, J_3,Fꢄ 17.6 Hz), 66.69 (C-4), 60.57 (C-

6), 41.69, 37.13, 30.21 (CH₂).

Anal. Calcd for C₂₄H₂₈FO₆N: C, 64.70; H, 6.34; N, 3.14. Found: C, 64.59; H,

6.37; N, 2.86.

Poly[p-[2-[N-(p-vinylbenzyl)carbamoyl]ethyl]phenyl ꢀ-D-mannopyra-

noside] (30). To a solution of 26 (269 mg, 0.61 mmol) in DMSO (1.0 mL), 2,2ꢂ-

azobisisobutyronitrile (4.0 mg, 0.02 mmol) was added. The reaction mixture was

set in a sealed tube, degassed with argon, and kept at 65°C in an oil bath for 24 h.

The reaction mixture was poured into stirred acetone, and the precipitate separated

using a glass filter to give 30 (260 mg, 97% yield): IR (cm^ꢃ1) 3340 (OH), 1644

(CꢄO); ¹³C NMR (A-500, d₆-DMSO) ꢈ 171.24 (CꢄO), 128.58, 115.76 (Ph),

126.58 (-CH-CH₂-), 97.78 (C-1), 77.35 (C-2), 73.41 (C-3), 70.45 (C-4), 66.77 (C-

5), 60.99 (C-6), 41.70, 40.40, 37.24, 30.51 (CH₂).

Poly[p-[2-[N-(p-vinylbenzyl)carbamoyl]ethyl]phenyl 2-acetamido-2-de-

oxy-ꢀ-D-mannopyranoside] (31). To a solution of 27 (204 mg, 0.42 mmol) in

DMSO (1.0 mL), 2,2ꢂ-azobisisobutyronitrile (4.0 mg, 0.02 mmol) was added. The

reaction mixture was set in a sealed tube, degassed with argon, and kept at 65°C in

an oil bath for 24 h. The reaction mixture was poured into stirred acetone, and the

precipitate separated using a glass filter to give 31 (171 mg, 84% yield): IR (cm^ꢃ1

)

3430 (OH), 1644 (CꢄO); ¹³C NMR (A-500, d₆-DMSO) ꢈ 170.19 (CꢄO), 128.95,

116.06 (Ph), 126.58 (-CH-CH₂-), 96.85 (C-1), 77.75 (C-5), 71.97 (C-3), 66.91 (C-

4), 61.02 (C-6), 52.38 (C-2), 37.22, 30.63, 30.33, 26.77 (CH₂), 22.92 (CH₃).

Poly[p-[2-(N-(p-vinylbenzyl)carbamoyl)ethyl]phenyl ꢀ-D-arabino-hex-

opyranoside] (32). To a solution of 28 (234 mg, 0.55 mmol) in DMSO (2.0 mL),

2,2ꢂ-azobisisobutyronitrile (3.0 mg, 0.015 mmol) was added. The reaction mixture

was set in a sealed tube, degassed with argon, and kept at 65°C in an oil bath for

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141

24 h. The reaction mixture was poured into stirred acetone, and the precipitate sep-

arated using a glass filter to give 32 (202 mg, 86% yield): IR (cm^ꢃ1) 3412 (OH),

1647 (CꢄO); ¹³C NMR (A-500, d₆-DMSO) ꢈ 171.48 (CꢄO), 128.95, 115.87 (Ph),

126.36 (-CH-CH₂-), 96.73 (C-1), 77.10 (C-5), 71.18 (C-), 70.39 (C-4), 60.80 (C-

6), 39.34 (C-2), 37.27, 30.63, 30.33 (CH₂).

Poly[p-[2-[N-(p-vinylbenzyl)carbamoyl]ethyl]phenyl 2-deoxy-2-fluoro-

ꢀ-D-mannopyranoside] (33). To a solution of 29 (229 mg, 0.514 mmol) in

DMSO (1.0 mL), 2,2ꢂ-azobisisobutyronitrile (4.0 mg, 0.02 mmol) was added. The

reaction mixture was set in a sealed tube, degassed with argon, and kept at 65°C in

an oil bath for 24 h. The reaction mixture was poured into stirred acetone, and the

precipitate separated using a glass filter to give 33 (128 mg, 56% yield): IR (cm^ꢃ1

)

3448 (OH), 1638 (CꢄO); ¹³C NMR (A-500, d₆-DMSO) ꢈ 171.37 (CꢄO), 129.03,

115.72 (Ph), 127.76 (-CH-CH₂-), 95.71 (C-1, J_1,Fꢄ 14.5 Hz), 90.51 (C-2, J_2,Fꢄ

184.1 Hz), 77.40 (C-5), 71.76 (C-3, J_3,Fꢄ 18.6 Hz), 66.68 (C-4), 60.58 (C-6),

40.09, 37.25, 30.65, 30.30 (CH₂).

Benzyl 3-(p-hydroxyphenyl)propionate (34). To a solution of 3-(p-hy-

droxyphenyl)propionic acid (500 mg, 3.0 mmol) and potassium hydrogencarbon-

ate (600 mg, 6.0 mmol) in N,N-dimethylformamide (10 mL), benzyl bromide (0.4

mL, 3.39 mmol) was added dropwise at 0°C and the reaction mixture was stirred

for 12 h at rt until the disappearance of starting compound on TLC with

hexane/ethyl acetate (4/1 v/v). The reaction mixture was poured into a saturated aq

NaHCO₃solution, extracted with ethyl acetate, washed with brine and water, dried

over anhydrous magnesium sulfate, filtered, and concentrated to give a residue.

The remaining residue was purified on a column of silica gel with hexane/ethyl ac-

etate (4/1 v/v). The yield of 34 was quantitative (765 mg): IR (cm^ꢃ1) 3406 (OH),

1713 (CꢄO); ¹H NMR (EX-90, CDCl₃) ꢈ 7.34–7.32 (m, 5H, -CH₂Ph), 7.04, 6.73

(each d, 4H, phenylene, J ꢄ 8.6 Hz), 5.33 (m, 1H, OH), 5.10 (s, 2H, -CH₂Ph), 2.85

(t, 2H, -CH₂CH₂CO-, J ꢄ 7.8 Hz), 2.68 (t, 2H, -CH₂CH₂CO-).

Anal. Calcd for C₁₆H₁₆O₃: C, 74.98%; H, 6.29%. Found: C, 74.49%; H,

6.44%.

Enzyme-linked lectin assay (ELLA)^23–25using Concanavaline A and the

synthetic glycopolymers as inhibitors. Nunclon microtiter plates (Nuc439454)

were coated with soybean lectin (SBA) (Honen Co. Ltd.: 03907) at 100 ꢇL/well of

2-fold serial dilution in 50 ꢇg/L Tris-HCl buffer (pH 7.3), containing 0.15 M NaCl,

and 0.1% (v/v) Tween 20 (PBST) and incubated overnight at 4°C. The plates were

then washed with 50 mM Tris-HCl buffer, containing 0.15M NaCl, and 0.1%

Tween 20, and blocked with 400 ꢇL of 1% (v/v) BSA in PBS for 1 h at rt and then

the plate was washed three times. Each inhibitor, methyl ꢁ-D-mannopyranoside,

methyl ꢁ-D-glucopyranoside, and synthesized glycopolymers (6, 30, 31, 32, 33)

(50% (v/v) DMSO/H₂O), was added to biotin labeled-Con A (Honen Co. Ltd.:

90408) (10 ng/mL in washing solution containing 0.25% (v/v) gelatine). After stor-

ing for 60 min at rt, 100 ꢇL of the above inhibitor solutions were transferred to the

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AKAI ET AL.

SBA-coated plates and incubated for 60 min at rt. After washing three times as

mentioned above, 100 ꢇL of horseradish peroxidase labeled-streptavidin (Fu-

nakoshi Co. Ltd.: Vector SA-5004) (1 ꢇg/mL in washing solution) was added, and

then incubated for 30 min at rt. After washing three times as mentioned above, 100

ꢇL of color-producing reagent (Funakoshi Co. Ltd.: 50507600) was added. After

30 min at rt, the reaction was stopped with 100 ꢇL of 2 mM NaN₃solution, and the

adsorbance of each well at 450 relative to 600 nm was determined on a micro plate

reader (Tosho Co. Ltd.: MPR-A4i).

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Received September 19, 2000

Accepted January 22, 2001

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Synthesis of new glycopolymers containing β-d-mannopyranose, and c-2-substituted β-d-mannopyranose residues as a new class of inhibitor

DOI: 10.1081/CAR-100103953

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Article abstract of DOI:10.1081/CAR-100103953

Full text of DOI:10.1081/CAR-100103953

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