Synthesis of β-D-Galp-(1→4)-β-D-GlcpNAc-(1→2)-α -D-Manp-(1→O)(CH2)7CH3 mimics to explore the substrate specificity of sialyltransferases and trans-sialidases

FULL PAPER

Synthesis of β-

D

-Galp-(1Ǟ4)-β-

D

-GlcpNAc-(1Ǟ2)-α-

D

-Manp-

(1ǞO)(CH₂)₇CH₃Mimics to Explore the Substrate Specificity of

Sialyltransferases and trans-Sialidases

John A. F. Joosten,^[a]Britta Evers,^[a]Ruben P. van Summeren,^[a]Johannis P. Kamerling,*^[a]

and Johannes F. G. Vliegenthart^[a]

Keywords: Carbohydrates / Glycosylations / Oligosaccharides / Sialyl transfer

Eleven trisaccharide octyl glycosides related to the N-glycan

sequence β- -Galp-(1Ǟ4)-β- -GlcpNAc-(1Ǟ2)-α- -Manp-

(1ǞO)(CH₂)₇CH₃(1) designed for detailed exploration of the

acceptor specificity of α-2,3- and α-2,6-sialyltransferases as

(CH₂)₇CH₃(9, R = R¹= Ac; 10, R = R¹= Pr; 11, R = Ac, R¹

=

D

Pr; 12, R = Pr, R¹= Ac). All trisaccharides were obtained by

condensation of suitably modified glycosyl donors based on

imidates or thioglycosides with the single disaccharide ac-

well as trans-sialidases, have been synthesised: β-

(1Ǟ4)-β- -GlcpNPr-(1Ǟ2)-α- -Manp-(1ǞO)(CH₂)₇CH₃(2), glucopyranosyl)-(1Ǟ2)-3,4,6-tri-O-benzyl-α-

β- -Fucp-(1Ǟ4)-β- -GlcpNR-(1Ǟ2)-α-

(CH₂)₇CH₃(3, R = Ac; 4, R = Pr), 6-amino-6-deoxy-β-

(1Ǟ4)-β- -GlcpNR-(1Ǟ2)-α- -Manp-(1ǞO)(CH₂)₇CH₃

R = Ac; 6, R = Pr), 2-deoxy-β- -Galp-(1Ǟ4)-β- -GlcpNR-

(1Ǟ2)-α- -Manp-(1ǞO)(CH₂)₇CH₃(7, R = Ac; 8, R = Pr), β-

-GalpNR¹-(1Ǟ4)-β-

-GlcpNR-(1Ǟ2)-α- -Manp-(1ǞO)-

D

-Galp-

ceptor octyl (3,6-di-O-benzyl-2-deoxy-2-phthalimido-β-

D-

D

-manno-

D

-Manp-(1ǞO)-

-Galp-

(5,

pyranoside, followed by deprotection. For the trisaccharides

containing an N-acylated glucosamine as well as an N-acyl-

ated galactosamine unit, use was made of a combination of

N-phthaloyl and N-dimethylmaleoyl protection.

( Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim,

Germany, 2003)

D

Introduction

ics, mainly deoxygenated forms of the pyranose rings, were

studied.^[6Ϫ8]It turned out that both the HO-6 group of the

galactose residue and the N-acetyl group of the N-acetylglu-

cosamine unit are required for the activity of α-2,6-sialyl-

Sialic acids occur at the non-reducing termini of many

glycoconjugates, and are considered to be key determinants

in the regulation of a variety of biological processes.^[1Ϫ3]In

the biosynthesis of human and animal sialylated glycans at

least 18 sialyltransferases are active with different substrate

specificities.^[2,4]Furthermore, several protozoal and bac-

terial sialidases have shown to act as trans-sialidases, trans-

transferase

I

(ST6Gal I). α-2,3-Sialyltransferase III

(ST3Gal III) required the HO-3, HO-4, and HO-6 groups

of the terminal galactose residue, and some influence from

the subterminal N-acetylglucosamine was noticed.

Currently, we have studied sialyltransferases involved in

ferring sialyl residues in α-2,3- or α-2,6-linkage from one the α-2,3- and α-2,6-sialylation of terminal galactose units

glycan to the terminal galactose unit of another non-sialyl- in N-glycoprotein glycans. At first instance it was shown

ated oligosaccharide or glycoconjugate.^[2,5]One of the most that rat liver ST6Gal I recognises, in addition to β--Galp-

abundant elements that is sialylated is the β--Galp-(1Ǟ4)- (1Ǟ4)-β--GlcpNAc-(1ǞOMe) and β--GalpNAc-(1Ǟ4)-

β--GlcpNAc-(1Ǟ2)-α--Manp-(1Ǟ sequence in glyco- β--GlcpNAc-(1ǞOMe),

also

β--Manp-(1Ǟ4)-β--

protein N-glycans. A thorough analysis of the substrate GlcpNAc-(1ǞOMe).^[9]Then, the trisaccharide β--Galp-

specificity of sialyltransferases and trans-sialidases can be (1Ǟ4)-β--GlcpNAc-(1Ǟ2)-α--Manp-(1ǞO)(CH₂)₇CH₃

attained by using modified oligosaccharides, probing the and eleven analogues containing structural variants of -

contribution of individual hydroxy groups and the N-acetyl- galactose were synthesised and employed as substrates for

ated amino function in recognition and binding.

rat liver ST6Gal I, recombinant full length human liver

Initially, focusing on identifying key polar groups re- ST6Gal I, and recombinant N-terminal truncated rat liver

quired for transfer of sialic acid from CPM-Neu5Ac by rat ST3Gal III.^[10,11,12]Hydroxy groups at either C-3 or C-4 of

liver sialyltransferases, the acceptors β--Galp-(1Ǟ3)-β-- the -galactose residue were substituted by hydrogen (3-

GlcpNAc and β--Galp-(1Ǟ4)-β--GlcpNAc, using mim- and 4-deoxy-β--Galp-R) or fluorine (3- and 4-fluoro-β--

Galp-R), by amino (3-amino-β--Galp-R) or O-methyl

[a]

groups (3- and 4-O-methyl-β--Galp-R), or were inverted

(β--Gulp-R and β--Glcp-R), to determine their involve-

ment in binding to and catalytic activity of the enzyme. In

addition, trisaccharides containing α--Altp (inverted

Bijvoet Center, Department of Bio-Organic Chemistry, Section

of Glycoscience and Biocatalysis, Utrecht University,

Padualaan 8, 3584 CH Utrecht, The Netherlands

Fax: (internat.) ϩ 31-30/2540980

E-mail: j.p.kamerling@chem.uu.nl

Eur. J. Org. Chem. 2003, 3569Ϫ3586

DOI: 10.1002/ejoc.200300293

 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

3569

J. P. Kamerling et al.

FULL PAPER

hydroxymethyl group at C-5) or β--Galp (enantiomer) at tyl) for 1, 3, 5, 7, 9, and 11, and R ϭ Pr (N-propionyl) for

the non-reducing terminus were constructed as probes. The 2, 4, 6, 8, 10, and 12, the key disaccharide acceptor was the

ST6Gal I tolerated most of the modifications at the -ga- earlier synthesised octyl (3,6-di-O-benzyl-2-deoxy-2-phthal-

lactose residue to some extent. The best substrates were the imido-β--glucopyranosyl)-(1Ǟ2)-3,4,6-tri-O-benzyl-α--

4-deoxy and 4-fluoro analogues, followed by the 3-deoxy mannopyranoside (13),^[10]having a free HO-4Ј function for

and 3-fluoro analogues. The ST3Gal III displayed a nar- elongation with suitably modified glycosyl residues. The fi-

rower specificity; only the 4-O-methyl analogue showed to nal glycosyl residues incorporated were mimics of the native

be a relatively good substrate, and the 4-deoxy and 4-fluoro β--galactopyranosyl residue, i.e. a 6-deoxy-β--galactopy-

analogues show a minor activity.^[13]Recently, it was demon- ranosyl (β--fucopyranosyl; 3 and 4), a 6-amino-6-deoxy-

strated that also conformationally constrained oligosacch- β--galactopyranosyl (5 and 6), a 2-deoxy-β--lyxo-hexo-

arides can act as acceptors for rat liver ST6Gal I.^[14]

pyranosyl (2-deoxy-β--galactopyranosyl; 7 and 8), a 2-acet-

In the framework of ongoing studies focused on the amido-2-deoxy-β--galactopyranosyl (9 and 12), and a 2-

understanding of the substrate specificity of sialyltransfer- deoxy-2-propionamido-β--galactopyranosyl (10 and 11)

ases and trans-sialidases, we have synthesised an additional residue.

series of 11 trisaccharide octyl glycosides. The trisacchar-

ides are of the type β--Sugp-(1Ǟ4)-β--GlcpNR-(1Ǟ2)-α-

Compounds 1 and 2

Trisaccharide 1, octyl β--galactopyranosyl-(1Ǟ4)-2-acet-

amido-2-deoxy-β--glucopyranosyl-(1Ǟ2)-α--manno-

pyranoside (Scheme 1), was synthesised as described earlier,

starting by coupling 2,3,4,6-tetra-O-acetyl-α--galactopyr-

anosyl trichloroacetimidate (14)^[15]with acceptor 13, using

trimethylsilyl trifluoromethanesulfonate as a catalyst, to

yield trisaccharide derivative 15^[10](Scheme 2). Along a

similar route octyl β--galactopyranosyl-(1Ǟ4)-2-deoxy-2-

propionamido-β--glucopyranosyl-(1Ǟ2)-α--manno-

pyranoside (2) (Scheme 1), the N-propionylated analogue of

-Manp-(1ǞO)(CH₂)₇CH₃, with modifications at C-6

(Sug ϭ 6-deoxy-Gal or 6-amino-6-deoxy-Gal) or at C-2

(Sug ϭ 2-deoxy-Gal, GalNAc or GalNPr) of the -galac-

tose residue, in combination with N-acetylated or N-propi-

onylated -glucosamine [R ϭ Ac or Pr (note that Pr ϭ pro-

pionyl and not propyl throughout the paper)].

Results and Discussion

In the synthesis of the trisaccharide variants 1Ϫ12 1, was prepared. To this end 15 was de-O-acetylated, then

(Scheme 1), all containing the element Ǟ4)-β--GlcpNR- de-N-phthaloylated using 1,2-diaminoethane in 1-butanol

(1Ǟ2)-α--Manp-(1ǞO)(CH₂)₇CH₃with R ϭ Ac (N-ace- at 90 °C^[16](Scheme 2), and N,O-propionylated using propi-

Scheme 1. List of structures of synthesized trisaccharide octyl glycosides

3570

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Eur. J. Org. Chem. 2003, 3569Ϫ3586

β-D-Galp-(1Ǟ4)-β-D-GlcpNAc-(1Ǟ2)-α-D-Manp-(1ǞO)(CH₂)₇CH₃Mimics

FULL PAPER

Scheme 2. Synthesis of trisaccharide 2: a) NaOMe (pH ϭ 9), CH₂Cl₂, MeOH; b) NH₂CH₂CH₂NH₂, 90 °C, 1-BuOH, molecular sieves

˚

(3 A); c) Pr₂O, pyridine; d) NaOMe (pH ϭ 9), MeOH; e) Ac₂O, pyridine, 72% over five steps; f) 10% Pd/C, H₂C, HOAc, EtOH, EtOAc;

g) Ac₂O, pyridine, 65% over two steps; h) NaOMe (pH ϭ 9), CH₂Cl₂, MeOH, 94%

onic anhydride in pyridine. Subsequent de-O-propi- benzylation, de-O-propionylation, and O-acetylation (Ǟ

onylation and O-acetylation (acetic anhydride in pyridine) 24, 55%), and de-O-acetylation yielded 4 (93%) (Scheme 4).

yielded 16 in an overall yield of 72%. Then, de-O-benzyl- The last O-acetylation step was carried out to facilitate

1

ation of 16 by hydrogenation using 10% Pd/C as a catalyst, chromatographic purification. The H and ¹³C NMR spec-

followed by O-acetylation (Ǟ 17, 65%) and finally de-O- troscopic data of 3 and 4 are presented in Tables 1 and 2,

acetylation yielded 2 (94%). The last O-acetylation step was respectively.

carried out to facilitate chromatographic purification. The

¹H and ¹³C NMR spectroscopic data of 1 and 2 are pre-

Compounds 5 and 6

sented in Tables 1 and 2, respectively.

For the syntheses of the 6ЈЈ-amino-6ЈЈ-deoxy mimics of 1

and 2, octyl 6-amino-6-deoxy-β--galactopyranosyl-(1Ǟ4)-

2-acetamido-2-deoxy-β--glucopyranosyl-(1Ǟ2)-α--man-

Compounds 3 and 4

For the syntheses of the 6ЈЈ-deoxy mimics of 1 and 2, nopyranoside (5) and octyl 6-amino-6-deoxy-β--galactopyr-

octyl β--fucopyranosyl-(1Ǟ4)-2-acetamido-2-deoxy-β-- anosyl-(1Ǟ4)-2-deoxy-2-propionamido-β--glucopyr-

glucopyranosyl-(1Ǟ2)-α--mannopyranoside (3) and octyl anosyl-(1Ǟ2)-α--mannopyranoside

(6)

(Scheme 1),

β--fucopyranosyl-(1Ǟ4)-2-deoxy-2-propionamido-β-- respectively, the galactosyl imidate donor 30 with an azide

glucopyranosyl-(1Ǟ2)-α--mannopyranoside (4) (Scheme 1), function at C-6 was applied (Scheme 5). This donor was

respectively, ethyl 2,3,4-tri-O-acetyl-1-thio-β--fucopyrano- synthesised starting from 1,2:3,4-di-O-isopropylidene-α--

side (19) was used as a donor (Scheme 3). To this end - galactopyranose (25). Treatment of 25 with p-toluenesul-

fucose 18 was O-acetylated, after which the thioethyl func- fonyl chloride in pyridine (Ǟ 26, 92%) and subsequent dis-

tion was introduced by reaction with ethanethiol in the placement of the tosyl group by an azide group using so-

presence of boron trifluorideϪdiethyl ether. Separation of dium azide in dimethyl sulfoxide yielded 27 (99%). The in-

the anomeric mixture by column chromatography yielded troduction of the azide group in 27 was verified by IR spec-

the β-anomer 19 in a yield of 61%, whereas the α-anomer troscopy [ν˜

ϭ

2110 cm^Ϫ1]. Then, 27 was de-O-

was isolated in a yield of 36%. Donor 19 was coupled with isopropylidenated using aqueous trifluoroacetic acid and O-

13 by in situ activation of the donor with bromine/silver acetylated (Ǟ 28, 50%), followed by selective de-O-acety-

trifluoromethanesulfonate, yielding trisaccharide derivative lation at C-1 using hydrazinium acetate in dimethylformam-

20 (74%) (Scheme 3). It should be noted that fucosylation ide (Ǟ 29, 89%), and imidation with trichloroacetonitrile in

using 2,3,4-tri-O-acetyl--fucopyranosyl trichloroacetimid- the presence of 1,8-diazabicyclo[5.4.0]undec-7-ene, to yield

ate was not successful (data not shown). De-N-phthaloyl- 30 (63%). Donor 30 was coupled with 13, using trimethyl-

ation/de-O-acetylation of 20, followed by N,O-acetylation silyl trifluoromethanesulfonate as a catalyst, affording tri-

(Ǟ 21, 85%), de-O-benzylation, and O-acetylation yielded saccharide derivative 31 (61%) (Scheme 6). De-N-phthaloyl-

22 (72%). Finally, de-O-acetylation of 22 afforded 3 (61%). ation/de-O-acetylation and N,O-acetylation of 31 (Ǟ 32,

In a similar way, de-N-phthaloylation/de-O-acetylation of 99%), followed by de-O-acetylation (Ǟ 33, 70%) and re-

20, followed by N,O-propionylation (Ǟ 23, 87%), de-O- duction with hydrogen in the presence of 10% Pd/C of the

Eur. J. Org. Chem. 2003, 3569Ϫ3586

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 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

3571

J. P. Kamerling et al.

FULL PAPER

1

Table 1. 500 MHz H NMR chemical shifts of the carbohydrate residues in the compounds 1Ϫ12

Residue^[a]

Man

Reporter group

1

2

3

4

H-1

4.858 (1.4)

4.038 (3.2)

3.80

4.849 (1.5)

4.048 (3.5)

3.79

4.860 (1.6)

4.041 (3.3)

3.79

4.808

4.006

3.79

H-2

H-3

H-4

3.53

3.50

3.51

H-5

3.60

3.59

3.58

3.51

H-6a^[b]

H-6b^[b]

H-1

3.63

3.62

3.63

3.68

3.89

3.88

3.79

GlcNHR^[c]

4.583 (7.0)

3.75

4.597 (7.6)

3.75

4.583 (7.6)

3.75

4.609 (7.6)

3.79

H-2

H-3

3.73

3.74

3.72

3.75

H-4

3.76

3.71

3.68

H-5

3.56

3.57

3.58

H-6a

H-6b

CH₂

CH₃

H-1

3.98

3.97

3.96

3.84

3.82

3.84

Ϫ

2.323

1.138

4.470 (7.7)

3.55

Ϫ

2.324

1.142

4.429 (7.8)

3.51

2.052

4.470 (7.7)

3.55

2.052

4.423 (7.9)

3.51

Gal

H-2

H-3

3.67

3.66

3.67

H-4

3.93 (3.2^[d]

3.74

)

3.92 (3.3^[d]

3.74

)

3.75

3.76

H-5

3.84

H-6a

H-6b

OCHH

CH₃

3.75

1.252 (6.4^[e]

Ϫ

)

1.257 (6.5^[e]

Ϫ

)

3.75

Spacer

3.53

3.52

3.53

3.48

3.74

3.79

3.72

3.69

0.872

0.861

0.868

Residue

Man

Reporter group

5

6

7

8

H-1

4.855

4.032

3.80

4.845

4.043

3.81

4.854 (1.5)

4.033 (3.3)

3.78

4.820

4.018

3.78

H-2

H-3

H-4

3.55

3.51

3.57

H-5

3.56

3.59

3.58

3.54

H-6a^[b]

H-6b^[b]

H-1

3.65

3.62

3.63

3.65

3.86

3.87

3.82

GlcNHR^[c]

4.585 (7.8)

3.75

4.596 (8.1)

3.77

4.569 (8.1)

3.75

4.589 (7.8)

3.75

H-2

H-3

3.74

3.73

3.72

H-4

3.78

3.76

3.75

H-5

3.55

3.51

H-6a

H-6b

CH₂

CH₃

H-1

3.99

3.77^[b]

3.89^[b]

Ϫ

3.77^[b]

3.88^[b]

2.325

1.140

4.719

1.70

3.86

3.85

Ϫ

2.327

1.139

4.497 (7.7)

3.53

2.057

4.501 (7.6)

3.55

2.051

4.715

1.69

Gal

H-2_ax

H-2_eq

H-3

H-4

H-5

H-6a

H-6b

OCHH

CH₃

Ϫ

2.08

3.68

3.69

3.89

3.90

3.93

3.77

3.84

3.81

3.61

3.23

3.22

3.77

3.15

3.77

Spacer

3.51

3.50

3.75

3.71

3.73

3.71

0.865

0.861

0.865

0.867

Residue

Man

Reporter group

9

10

11

12

H-1

4.852

4.026

3.79

4.840

4.027

3.78

4.846

4.008

3.79

4.837

4.027

3.79

H-2

H-3

H-4

3.50

3.51

3.55

3.50

H-5

3.58

3.59

3.56

3.57

H-6a^[b]

H-6b^[b]

H-1

3.62

3.61

3.64

3.60

3.88

3.87

3.85

3.86

GlcNHR¹

4.558 (7.9)

3.73

4.565 (7.9)

3.73

4.551 (8.0)

3.71

4.571 (7.5)

3.73

[f]

H-2

H-3

3.72

3.74

3.75

3.74

H-4

3.63

3.64

3.68

3.64

H-5

3.47

3.48

3.46

3.48

H-6a^[b]

H-6b^[b]

CH₂

CH₃

H-1

3.66

3.67

3.68

3.66

3.84

3.85

3.84

Ϫ

2.32

Ϫ

2.32

2.045

4.516 (8.4)

3.94

1.133

4.517 (8.4)

3.94

2.047

4.523 (8.4)

3.95

1.133

4.519 (8.3)

3.92

GalNHR^2[f]

H-2

H-3

3.78

3.75

3.76

H-4

3.93

3.94

H-5

3.72

3.73

3.75

3.73

H-6a

H-6b

CH₂

CH₃

OCHH

CH₃

3.78

3.81

3.78

3.81

Ϫ

2.32

2.33

Ϫ

2.067

1.133

3.55

1.133

3.50

2.068

Spacer

[a]

3.53

3.51

3.73

3.75

3.71

3.73

0.861

0.860

0.866

0.861

Data were measured in D₂O at 300 K. Chemical shifts are relative to internal acetone (δ ϭ 2.225 ppm). Coupling constants are given

[b]

[c]

in Hertz between parentheses.

odd numbers and propionyl for even numbers.

for 9 and 11, and propionyl for 10 and 12. R²ϭ acetyl for 9 and 12, and propionyl for 10 and 11.

The assignment of H-6a and H-6b may have to be interchanged within one residue. R ϭ acetyl for

J_3ЈЈ,4ЈЈin 1 and 2. H-6,6,6 signal and J_5ЈЈ,6ЈЈfor -fucose in 3 and 4. R¹ϭ acetyl

[d]

[e]

[f]

3572

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Eur. J. Org. Chem. 2003, 3569Ϫ3586

β-D-Galp-(1Ǟ4)-β-D-GlcpNAc-(1Ǟ2)-α-D-Manp-(1ǞO)(CH₂)₇CH₃Mimics

FULL PAPER

Table 2. 125 MHz ¹³C NMR chemical shifts of the carbohydrate

Table 2. (Continued)

residues in the compounds 1Ϫ12

Residue^[a]

Man

Reporter group

1

2

3

4

Residue

Man

Reporter group

9

10

11

12

C-1

C-2

C-3

C-4

C-5

C-6

C-1

C-2

C-3

C-4

C-5

C-6

CO

CH₂

CH₃

C-1

C-2

C-3

C-4

C-5

C-6

OCH₂

CH₂

CH₃

97.5

77.3

70.4

67.8

73.4

62.0

100.1

55.6

72.5

79.2

75.3

60.6

175.2

Ϫ

22.9

103.5

71.5

73.1

69.1

75.9

61.6

68.7

31.7

29.2

29.1

29.0

26.0

22.6

14.0

97.5

77.4

70.4

68.0

73.5

62.1

100.2

55.5

72.7

79.3

75.4

60.7

179.0

30.0

9.9

103.6

71.6

73.2

69.2

76.0

61.6

68.7

31.7

29.1

29.0

28.9

26.0

22.6

14.0

97.4

77.3

70.4

67.9

73.5

62.1

100.1

55.5

72.6

79.8

75.3

60.6

175.2

Ϫ

22.9

103.6

71.3

73.3

71.8

71.7

15.9

68.7

31.7

29.1

29.0

28.9

26.0

22.6

14.0

98.0

78.3

70.6

67.5

73.5

61.8

100.8

55.6

72.5

79.8

75.2

60.7

178.5

29.9

9.9

103.6

71.3

73.4

71.8

71.7

16.0

68.5

32.1

29.6

29.5

29.4

26.3

22.9

14.2

C-1

C-2

C-3

C-4

C-5

C-6

97.4

77.3

70.4

67.9

73.5

62.1

97.4

77.3

70.4

67.9

73.5

62.1

97.4

77.3

70.4

67.7

73.5

62.0

97.5

77.4

70.4

67.9

73.5

62.0

100.2

55.3

72.8

79.9

75.1

60.7

179.0

29.9

9.9

102.4

53.2

71.3

68.2

75.9

61.5

175.3

Ϫ

GlcNHR^[b]

GlcNHR^{1 [c]}C-1

100.1 100.1 100.1

C-2

C-3

C-4

C-5

C-6

CO

CH₂

CH₃

C-1

C-2

C-3

C-4

55.4

72.7

79.9

75.1

60.7

55.2

72.8

79.6

75.2

60.8

55.4

72.6

79.5

75.2

60.7

175.2 179.0 175.2

Ϫ

23.0

30.0

9.9

Ϫ

23.0

Gal

GalNHR^2[c]

102.4 102.2 102.3

53.2

71.3

68.2

75.9

61.5

53.0

71.3

68.3

75.9

61.6

53.0

71.3

68.3

75.9

61.6

C-5

C-6

CO

Spacer

175.2 179.1 178.9

Ϫ

CH₂

CH₃

OCH₂

CH₂

CH₃

30.0

9.9

29.9

9.9

22.8

68.7

31.7

29.1

29.0

26.0

22.6

14.0

22.8

68.7

31.7

29.1

29.0

26.0

22.6

14.0

Spacer

68.7

31.7

29.1

29.0

28.9

26.0

22.6

14.0

68.6

31.9

29.4

29.3

29.2

26.2

22.7

14.1

Residue

Man

Reporter group

5

6

7

8

C-1

C-2

C-3

C-4

C-5

C-6

C-1

C-2

C-3

C-4

C-5

C-6

CO

CH₂

CH₃

C-1

C-2

C-3

C-4

C-5

C-6

OCH₂

CH₂

CH₃

97.5

77.5

70.4

67.8

73.5

62.0

100.2

55.8

72.5

77.5

75.5

60.4

175.3

Ϫ

23.0

103.1

71.5

73.0

69.7

72.7

41.1

68.7

31.8

29.3

29.2

29.0

26.1

22.6

14.0

97.6

77.7

70.4

67.9

73.5

62.0

100.4

55.7

72.5

77.6

75.5

60.4

179.0

29.9

9.9

103.1

71.5

73.0

69.8

72.9

41.1

68.7

31.7

29.1

29.0

26.0

22.6

14.0

97.5

77.3

70.4

67.8

73.5

62.1

100.2

55.6

72.6

79.1

75.3

60.7

175.2

Ϫ

23.0

101.1

34.1

68.3

67.3

76.2

62.0

68.7

31.8

29.2

29.0

26.1

22.6

14.0

97.8

77.9

70.5

67.6

73.5

62.0

100.6

55.7

72.6

79.1

75.3

60.8

178.7

29.9

9.9

101.1

34.2

68.3

67.3

76.2

61.9

68.5

31.9

29.4

29.2

26.2

22.8

14.1

[a]

Data measured in D₂O at 300 K. Chemical shifts are relative to

[b]

internal acetone (δ_CH3ϭ 30.89 ppm). R ϭ acetyl for odd num-

bers and propionyl for even numbers. R¹ϭ acetyl for 9 and 11,

[c]

and propionyl for 10 and 12. R²ϭ acetyl for 9 and 12, and propi-

onyl for 10 and 11.

GlcNHR^[b]

azido function (pH ϭ 10, ammonia; conversion into amino)

and of the O-benzyl groups (pH ഠ 5, acetic acid; conversion

into hydroxy), yielded 5 (99%). In a similar way, de-N-

phthaloylation/de-O-acetylation and N,O-propionylation of

31 (Ǟ 34, 94%), followed by de-O-propionylation (Ǟ 35,

79%), and conversion of the azido function into an amino

group combined with the removal of the O-benzyl groups,

yielded 6 (48%) (Scheme 7). The ¹H and ¹³C NMR spectro-

scopic data of 5 and 6 are presented in Tables 1 and 2,

respectively.

Gal

Spacer

Compounds 7 and 8

For the syntheses of the 2ЈЈ-deoxy mimics of 1 and 2,

octyl 2-deoxy-β--lyxo-hexopyranosyl-(1Ǟ4)-2-acetamido-

2-deoxy-β--glucopyranosyl-(1Ǟ2)-α--mannopyranoside

(7) and octyl 2-deoxy-β--lyxo-hexopyranosyl-(1Ǟ4)-2-de-

oxy-2-propionamido-β--glucopyranosyl-(1Ǟ2)-α--manno-

pyranoside (8) (Scheme 1), respectively, use was made of

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FULL PAPER

˚

Scheme 3. Synthesis of trisaccharide 3: a) Ac₂O, pyridine; b) 4 equiv. BF₃·Et₂O, 1.2 equiv. EtSH, 0 °C, CH₂Cl₂, molecular sieves (4 A),

˚

61% over two steps; c) 3.5 equiv. AgOTf, 1.1 equiv. Br₂, CH₂Cl₂, toluene, molecular sieves (4 A), 74%; d) NH₂CH₂CH₂NH₂, 90 °C, 1-

˚

BuOH, molecular sieves (3 A); e) Ac₂O, pyridine, 85% over two steps; f) 10% Pd/C, H₂, HOAc, EtOH, EtOAc; g) Ac₂O, pyridine, 72%

over two steps; h) NaOMe (pH ϭ 9), CH₂Cl₂, MeOH, 61%

˚

Scheme 4. Synthesis of trisaccharide 4: a) NH₂CH₂CH₂NH₂, 90 °C, 1-BuOH, molecular sieves (3 A); b) Pr₂O, pyridine, 87% over two

steps; c) 10% Pd/C, H₂, HOAc, EtOH, EtOAc; d) NaOMe (pH ϭ 9), MeOH; e) Ac₂O, pyridine, 55% over three steps; f) NaOMe (pH ϭ

9), MeOH, 93%

Scheme 5. Synthesis of 6-azido galactose donor 30: a) p-TosCl, CH₂Cl₂, pyridine, 92%; b) NaN₃, Me₂SO, 160 °C, 99%; c) TFA, H₂O,

CH₂Cl₂; d) Ac₂O, pyridine, 50% over two steps; e) hydrazinium acetate, DMF, 50 °C, 89%; f) Cl₃CCN, DBU, 0 °C, CH₂Cl₂, 63%

˚

Scheme 6. Synthesis of trisaccharide 5: a) 17% TMSOTf, CH₂Cl₂, 0 °C, molecular sieves (4 A), 61%; b) NH₂CH₂CH₂NH₂, 90 °C, 1-

˚

BuOH, molecular sieves (3 A); c) Ac₂O, pyridine, 99% over two steps; d) NaOMe (pH ϭ 9), MeOH, 70%; e) 10% Pd/C, H₂, NH₃, 2-

PrOH, H₂O, HOAc, 99%

3574

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β-D-Galp-(1Ǟ4)-β-D-GlcpNAc-(1Ǟ2)-α-D-Manp-(1ǞO)(CH₂)₇CH₃Mimics

FULL PAPER

˚

Scheme 7. Synthesis of trisaccharide 6: a) NH₂CH₂CH₂NH₂, 90 °C, 1-BuOH, molecular sieves (3 A); b) Pr₂O, pyridine, 94% over two

steps; c) NaOMe (pH ϭ 9), MeOH, 79%; d) 10% Pd/C, H₂, NH₃, 2-PrOH, H₂O, HOAc, 48%

ethyl 2-O-acetyl-3,4,6-tri-O-benzyl-1-thio-β--galactopyr- of catalytic amounts of α,αЈ-azoisobutyronitrile (AIBN) at

anoside (36).^[17]Coupling of 36 with 13, using N-iodosuccin- 100 °C^[18]afforded 41 (70%), which was catalytically de-O-

imide/trifluoromethanesulfonic acid as the catalytic sys- benzylated to give 7 (75%). In a similar way, de-N-

tem, yielded trisaccharide derivative 37 (70%) (Scheme 8). phthaloylation/de-O-acetylation and N,O-propionylation of

De-N-phthaloylation/de-O-acetylation and N,O-acetylation 37 (Ǟ 42, 94%), followed by de-O-propionylation (Ǟ 43),

of 37 (Ǟ 38, 94%), followed by de-O-acetylation (Ǟ 39) O-phenoxythiocarbonylation at C-2ЈЈ (Ǟ 44, 43%), 2ЈЈ-de-

and treatment with phenyl chlorothionocarbonate in 4-di- oxygenation (Ǟ 45, 62%), and de-O-benzylation yielded 8

methylaminopyridine/acetonitrile,^[18]yielded 40 (61%). 2ЈЈ- (57%) (Scheme 9). The ¹H and ¹³C NMR spectroscopic

Deoxygenation of 40 with tributylstannane in the presence data of 7 and 8 are presented in Tables 1 and 2, respectively.

˚

Scheme 8. Synthesis of trisaccharide 7: a) 1.4 equiv. NIS, 20% TfOH, CH₂Cl₂, 0 °C, molecular sieves (4 A), 70%; b) NH₂CH₂CH₂NH₂,

˚

90 °C, 1-BuOH, molecular sieves (3 A); c) Ac₂O, pyridine, 94% over two steps; d) NaOMe (pH ϭ 10), CH₂Cl₂, MeOH, 40 °C; e)

C₆H₅OC[S]Cl, DMAP, CH₃CN, 100 °C, 61% over two steps; f) Bu₃SnH, AIBN, 100 °C, toluene, 70%; g) 10% Pd/C, H₂, HOAc, EtOH,

EtOAc, 75%

˚

Scheme 9. Synthesis of trisaccharide 8: a) NH₂CH₂CH₂NH₂, 90 °C, 1-BuOH, molecular sieves (3 A); b) Pr₂O, pyridine, 94% over two

steps; c) NaOMe (pH ϭ 10), CH₂Cl₂, MeOH, 40 °C; d) C₆H₅OC[S]Cl, DMAP, CH₃CN, 100 °C, 43% over two steps; e) Bu₃SnH, AIBN,

100 °C, toluene, 62%; f) 10% Pd/C, H₂, HOAc, EtOH, EtOAc, 57%

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FULL PAPER

Compounds 9 and 10

CH₃,^[22]have been synthesised earlier along similar routes.

1

The H and ¹³C NMR spectroscopic data of 9 and 10 are

For the syntheses of the 2ЈЈ-acylamido-2ЈЈ-deoxy mimics

of 1 and 2, octyl 2-acetamido-2-deoxy-β--galactopyrano-

syl-(1Ǟ4)-2-acetamido-2-deoxy-β--glucopyranosyl-(1Ǟ2)-

α--mannopyranoside (9) and octyl 2-deoxy-2-propion-

amido-β--galactopyranosyl-(1Ǟ4)-2-deoxy-2-propion-

presented in Tables 1 and 2, respectively.

Compounds 11 and 12

The syntheses of the 2ЈЈ-acylamido-2ЈЈ-deoxy mimics of

amido-β--glucopyranosyl-(1Ǟ2)-α--mannopyranoside 1 and 2, octyl 2-deoxy-2-propionamido-β--galactopyrano-

(10) (Scheme 1), respectively, the earlier reported donor syl-(1Ǟ4)-2-acetamido-2-deoxy-β--glucopyranosyl-(1Ǟ2)-

ethyl 4-O-acetyl-3,6-di-O-benzyl-2-deoxy-2-phthalimido-1- α--mannopyranoside (11) and octyl 2-acetamido-2-deoxy-

thio-β--galactopyranoside (46)^[19]was used. This fragment β--galactopyranosyl-(1Ǟ4)-2-deoxy-2-propionamido-β--

was synthesised from a known glucosamine building glucopyranosyl-(1Ǟ2)-α--mannopyranoside

(12)

block,^[20]which was applied in the synthesis of disaccharide (Scheme 1), respectively, required in addition to the phthali-

13.^[10,11]Compound 46 was coupled to acceptor 13, using mido function for the glucosamine unit in acceptor 13, the

bromine/silver trifluoromethanesulfonate as the catalytic selection of another N-protective group for the galactosam-

system, to yield trisaccharide derivative 47 (51%) ine donor. Introductory experiments with donors containing

(Scheme 10). De-N-phthaloylation/de-O-acetylation and an N-acetyl or an N-(tetrachlorophthalimido)^[23,24]protective

N,O-acetylation of 47 (Ǟ 48, 61%), followed by de-O- group gave rise to irreproducible results only. Testing of the

benzylation and O-acetylation (Ǟ 49, 86%), and final de- promising N-(2,2,2-trichloroethoxycarbonyl) protective

O-acetylation yielded 9 (56%). In a similar way, de-N- group^[25Ϫ27]in terms of yields, showed problems in the de-

phthaloylation/de-O-acetylation and N,O-propionylation of protection using activated zinc;^[28]here also primary O-

47 (Ǟ 50, 80%), followed by de-O-benzylation and O-acety- benzyl groups were removed, leading to mixtures that could

lation (Ǟ 51, 92%), and final de-O-acylation yielded 10 not be fractionated. The N-(dimethylmaleoyl) protective

(91%) (Scheme 11). It should been noted that two ana- group^[29,30]turned out to be the best choice. According to

logues of 9, β--GalpNAc-(1Ǟ4)-β--GlcpNAc-(1Ǟ2)-α-- the earlier reported route for -glucosamine hydrochlo-

[21]

Manp-(1ǞO)(CH₂)₈COOCH₃

and

β--GalpNAc- ride,^[29,30]-galactosamine hydrochloride (52) was neutral-

(1Ǟ4)-β--GlcpNAc-(1Ǟ2)-α--Manp-(1ǞO)(CH₂)₂- ised with sodium methoxide in methanol, then N-acylated

˚

Scheme 10. Synthesis of trisaccharide 9: a) 0.5 equiv. Br₂, 1.2 equiv. AgOTf, CH₂Cl₂, toluene, molecular sieves (4 A), 51%; b)

˚

NH₂CH₂CH₂NH₂, 90 °C, 1-BuOH, molecular sieves (3 A); c) Ac₂O, pyridine, 61% over two steps; d) 10% Pd/C, H₂, HOAc, EtOH,

EtOAc; e) Ac₂O, pyridine, 86% over two steps; f) NaOMe (pH ϭ 10), MeOH, 56%

˚

Scheme 11. Synthesis of trisaccharide 10: a) NH₂CH₂CH₂NH₂, 90 °C, 1-BuOH, molecular sieves (3 A); b) Pr₂O, pyridine, 80% over two

steps; c) 10% Pd/C, H₂, HOAc, EtOH, EtOAc; d) Ac₂O, pyridine, 92% over two steps; e) NaOMe (pH ϭ 10), MeOH, 91%

3576

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β-D-Galp-(1Ǟ4)-β-D-GlcpNAc-(1Ǟ2)-α-D-Manp-(1ǞO)(CH₂)₇CH₃Mimics

FULL PAPER

Scheme 12. Synthesis of galactose donor 55: a) 1 equiv. NaOMe, DMMA, NEt₃, 60 °C, MeOH; b) Ac₂O, pyridine, 31% over two steps; c)

hydrazinium acetate, DMF, 61%; d) CCl₃CN, DBU, CH₂Cl₂, 59%

˚

Scheme 13. Synthesis of trisaccharide 11: a) 10% TMSOTf, 0 °C, CH₂Cl₂, molecular sieves (4 A), 84%; b) NaOH, HCl, dioxane, H₂O; c)

Pr₂O, pyridine, 33% over two steps; d) NH₂CH₂CH₂NH₂, 90 °C, 1-BuOH; e) Ac₂O, pyridine, 74% over two steps; f) NaOMe (pH ϭ 9),

CH₂Cl₂, MeOH; g) 10% Pd/C, H₂, HOAc, EtOH, EtOAc; h) Ac₂O, pyridine, 72% over three steps; i) NaOMe (pH ϭ 10), CH₂Cl₂,

MeOH, 77%

with dimethylmaleic anhydride in the presence of triethyl- acetylation of 56 by sequential incubation with sodium hy-

amine at 60 °C, and O-acetylated to give 53 (31%) droxide in aqueous dioxane (alkaline pH), and, after acidifi-

(Scheme 12). Subsequent de-O-acetylation at C-1 using hy- cation with HCl at pH ϭ 3,^[29,30]followed by N,O-propi-

drazinium acetate in dimethylformamide (Ǟ 54, 61%) and onylation yielded 57 (33%). De-N-phthaloylation/de-O-pro-

imidation with trichloroacetonitrile in the presence of 1,8- pionylation and N,O-acetylation of 57 (Ǟ 58, 74%), followed

diazabicyclo[5.4.0]undec-7-ene, yielded donor 55 (59%). by de-O-acetylation, de-O-benzylation, and O-acetylation

Coupling of 55 [N-(dimethylmaleoyl) function] with 13 (N- (Ǟ 59, 72%), and final de-O-acetylation yielded 11 (77%).

phthaloyl group), using trimethylsilyl trifluoromethanesul- The last O-acetylation step was carried out to facilitate chro-

fonate as a catalyst, afforded trisaccharide derivative 56 matographic purification. In

a

similar way, de-N-

(84%) (Scheme 13). De-N-(dimethylmaleoyl)ation and de-O- (dimethylmaleoyl)ation/de-O-acetylation and N,O-acety-

Scheme 14. Synthesis of trisaccharide 12: a) NaOH, HCl, AcCl, dioxane, H₂O; b) Ac₂O, pyridine, 48% over two steps; c) NH₂CH₂CH₂NH₂,

90 °C, 1-BuOH; d) Pr₂O, pyridine, 80% over two steps; e) NaOMe (pH ϭ 9), CH₂Cl₂, MeOH; f) 10% Pd/C, H₂, HOAc, EtOH, EtOAc; g)

Ac₂O, pyridine, 67% over three steps; h) NaOMe (pH ϭ 9), CH₂Cl₂, MeOH, 84%

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1

AMX 500 spectrometer. H NMR chemical shifts (δ_H) are given in

ppm relative to the signal for internal Me₄Si (δ_Hϭ 0 ppm) for solu-

tions in CDCl₃or by reference to acetone (δ_Hϭ 2.225 ppm) for

solutions in D₂O. ¹³C NMR chemical shifts (δ_C) are relative to the

signal for CDCl₃(δ_Cϭ 76.9 ppm) for solutions in CDCl₃or by

reference to acetone (δ_Cϭ 30.89 ppm) for solutions in D₂O. J values

are given in Hz. Optical rotations were determined for solutions in

CHCl₃or H₂O at 20 °C with a PerkinϪElmer 241 polarimeter, using

a 10-cm 1-mL cell. Fast-atom-bombardment mass spectrometry

(FABMS) was performed with a JEOL JMS SX/SX 102A four-sec-

lation of 56 (Ǟ 60, 48%) (Scheme 14), followed by de-N-

phthaloylation/de-O-acetylation and N,O-propionylation (Ǟ

61, 80%), and de-O-propionylation, de-O-benzylation, and

O-acetylation (Ǟ 62, 67%), and final de-O-acetylation

1

yielded 12 (84%). The H and ¹³C NMR spectroscopic data

of 11 and 12 are presented in Tables 1 and 2, respectively.

The NMR spectra of 11 and 12 are almost identical, except

for the N-acetyl methyl signal in the H NMR spectra. The

1

NAc signal in 11 resonates at δ ϭ 2.047 ppm, and in 12 at

δ ϭ 2.068 ppm. Note that in 9 the two NAc signals resonate tor mass spectrometer, operated at 10 kV accelerating voltage,

equipped with a JEOL MS-FAB 10 D FAB gun operated at 10 mA

emission current, producing a beam of 6 keV Xenon atoms.

MALDI-TOF mass spectra were recorded with a Voyager-DE (Per-

Septive Biosystems) instrument using dihydroxybenzoic acid (DHB)

as a matrix. The matrix was dissolved in a 1:1 (v/v) mixture of aceto-

nitrile/H₂O (10 mg DHB/mL), and the sample was dissolved in ace-

tone (5 mg/mL). Subsequently, 0.5 µL of matrix solution and 0.5 µL

of sample solution were brought on the sample plate of the mass

spectrometer. Spectra were generated by summing positive-ion sig-

nals of 256 laser shots with constant intensity. Exact masses were

measured by nano electrospray time-of-flight mass spectrometry

using a Micromass LCToF mass spectrometer at a resolution of 5000

FWHM. Gold-coated capillaries were loaded with 1 µL of sample

(conc. 20 µM), dissolved in a 1:1 (v/v) mixture of acetonitrile/H₂O

and 0.1% formic acid. Pentafluorophenylalanine was added as in-

ternal standard. The capillary voltage was set at 1500 V and the cone

voltage was set at 30 V.

at δ ϭ 2.045 and 2.067 ppm.

Conclusion

Summarising, to prepare a series of relevant trisaccharide

mimics for enzymatic sialyl-transfer studies, the strategy of

condensing a series of galactosyl-modified monosaccharide

donors with a general glucosaminyl-mannosyl acceptor has

proven to be successful. In earlier work, we have shown that

such an approach leads to better results than preparing di-

saccharide donors by introducing modifications at the galac-

tosyl-glucosaminyl level, followed by coupling to a fixed

mannosyl acceptor.^[12]In the present approach, monosac-

charide thioglycosides were used for coupling of synthons

containing besides acyl groups also benzyl groups or a deoxy

function (19, 36, 46), and trichloroacetimidates for coupling

of acylated deactivated synthons (14, 30, 55). For the acti-

vation of the thioglycosides, bromine/silver trifluoromethane-

sulfonate gave the best results for 19 and 46, whereby 19

needed more activation than 46, and N-iodosuccinimide/tri-

fluoromethanesulfonic acid for 36. TMSOTf gave always the

best results in trichloroacetimidate couplings.

Octyl (2,3,4,6-Tetra-O-acetyl-β-

O-benzyl-2-deoxy-2-propionamido-β-

tri-O-benzyl-α- -mannopyranoside (16): To

D

-galactopyranosyl)-(1Ǟ4)-(3,6-di-

-glucopyranosyl)-(1Ǟ2)-3,4,6-

solution of octyl

D

a

(2,3,4,6-tetra-O-acetyl-β--galactopyranosyl)-(1Ǟ4)-(3,6-di-O-

benzyl-2-deoxy-2-phthalimido-β--glucopyranosyl)-(1Ǟ2)-3,4,6-tri-

O-benzyl-α--mannopyranoside (15; 246 mg, 0.18 mmol)^[10]in

CH₂Cl₂/MeOH (20 mL, 3:1) was added NaOMe (pH ϭ 9), and the

mixture was stirred for 4 h. After neutralisation with Dowex 50 ϫ 8

(H^ϩ), and filtration, the solution was concentrated. A solution of

The trisaccharides 1Ϫ12 will be used in α-2,3- and α-2,6-

sialyltransferase as well as in trans-sialidase kinetic studies

in order to investigate the relevance in the substrate/enzyme

interaction of the replaced hydroxy functions in the terminal

galactose unit of 1 in combination with the effect of replac-

ing the N-acetyl function of the middle glucosamine unit by

an N-propionyl function.

˚

the residue in 1-BuOH (20 mL), containing molecular sieves 3 A

(0.75 g), was stirred for 30 min under Ar, then 1,2-diaminoethane

(2.2 mL, 33 mmol) was added. The mixture was stirred at 90 °C

overnight, filtered through Celite, and co-concentrated with toluene.

The residue was dissolved in pyridine/propionic anhydride (10 mL,

1:1) and stirred overnight, then co-concentrated with toluene. A

solution of the residue in MeOH (20 mL) was treated with NaOMe

(pH ϭ 9) for 4 h, then neutralised with Dowex 50 ϫ 8 (H^ϩ), filtered,

and concentrated. The residue was dissolved in pyridine/acetic anhy-

dride (10 mL, 1:1) and stirred overnight, then co-concentrated with

toluene. Column chromatography (toluene/EtOAc, 3:1) of the resi-

due yielded 16, isolated as a syrup (172 mg, 72%). TLC (toluene/

EtOAc, 1:1): R_fϭ 0.69. [α]²_D⁰ϭ ϩ3 (c ϭ 1, CHCl₃). ¹H NMR

(300 MHz, CDCl₃): δ ϭ 0.88 (t, 3 H, octyl CH₃), 1.00 (t, 3 H,

COCH₂CH₃), 1.26 (m, 10 H, 5 octyl CH₂), 1.50 (m, 2 H, octyl CH₂),

1.97, 1.98, 2.01, and 2.08 (4 s, each 3 H, 4 COCH₃), 2.15 (q, 2 H,

COCH₂CH₃), 3.13 and 3.42 (2 m, each 1 H, octyl OCH₂), 4.11 (d,

Experimental Section

General: All solvents used were distilled from appropriate drying ag-

ents. In the workup procedures of reaction mixtures, organic solu-

tions were washed with appropriate amounts of aqueous solutions

as indicated. Solutions were concentrated under reduced pressure at

40 °C (water bath). Reactions were monitored by TLC on Silica Gel

60 F₂₅₄(Merck), compounds were visualized under UV light, and

by charring with either 10% ethanolic H₂SO₄or 0.2% orcinol in

20% methanolic H₂SO₄. Column chromatography was performed on

Kieselgel 60 F₂₅₄(Merck, 0.063Ϫ0.200 mm). Size-exclusion chroma-

J_1,2Ͻ 1 Hz, 1 H, 1-H), 5.12 (dd, 1 H, 2ЈЈ-H), 5.17 (d, J_1Ј,2Јϭ 7.7 Hz,

1 H, 1Ј-H), 5.18 (d, J_2Ј,NHϭ 7.7 Hz, 1 H, NH), 5.28 (d, J_3ЈЈ,4ЈЈ

ϭ

1

tography was carried out on Sephadex LH-20 or Bio-Gel P-2. H

2.9, J_4ЈЈ,5ЈЈϽ 1 Hz, 1 H, 4ЈЈ-H), 7.45Ϫ7.10 (m, 25 H, 5 OCH₂C₆H₅)

ppm. MS (FAB^ϩ) of C₇₂H₉₁NO₂₀(1289): found m/z ϭ 1290 [M ϩ

H]^ϩ, 1312 [M ϩ Na]^ϩ.

(300 MHz) and ¹³C NMR spectra (75.5 MHz) of protected com-

pounds were recorded at 300 K using a Bruker AC 300 spectrometer;

only relevant NMR spectroscopic data are included in the exper-

imental procedures. Two-dimensional ¹H-¹H correlated spectra

Octyl (2,3,4,6-Tetra-O-acetyl-β-

D

-galactopyranosyl)-(1Ǟ4)-(3,6-di-

-glucopyranosyl)-(1Ǟ2)-3,4,6-

D-mannopyranoside (17): To a solution of 16 (172 mg,

(TOCSY, ROESY) and H-¹³C correlated spectra (HSQC) of com- O-acetyl-2-deoxy-2-propionamido-β-

D

1

pounds 1Ϫ12, 59 and 62 were recorded at 300 K using a Bruker tri-O-acetyl-α-

3578 www.eurjoc.org

 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

Eur. J. Org. Chem. 2003, 3569Ϫ3586

β-D-Galp-(1Ǟ4)-β-D-GlcpNAc-(1Ǟ2)-α-D-Manp-(1ǞO)(CH₂)₇CH₃Mimics

FULL PAPER

1.13 mmol) in EtOH/EtOAc (20 mL, 1:1) were added 10% Pd/C

(140 mg) and HOAc (0.25 mL), and the mixture was stirred for 2.5 h

under H₂, then filtered through Celite, and concentrated. A solution

of the residue in pyridine/acetic anhydride (22 mL, 1:1) was stirred

µmol) and 13 (250 mg, 242 µmol) in dry CH₂Cl₂(10 mL), containing

powdered molecular sieves 4 A (0.3 g), was stirred for 3.5 h under

Ar, then a solution of AgOTf (296 mg, 1.15 mmol) in dry toluene

(4.3 mL) was added, followed after 20 min by a solution of Br₂(371

˚

overnight, then co-concentrated with toluene. Column chromatogra- µmol) in dry CH₂Cl₂(0.69 mL). After additional stirring for 60 min,

phy (toluene/EtOAc, 1:3) of the residue yielded 17, isolated as a the mixture was neutralised with NEt₃and filtered through Celite.

syrup (88 mg, 65%). TLC (toluene/EtOAc, 1:3): R_fϭ 0.14. [α]_D²⁰

ϭ

The filtrate was diluted with CH₂Cl₂, washed with aq. 10% NaS₂O₃,

Ϫ7 (c ϭ 1, CHCl₃). ¹H NMR (300 MHz, CDCl₃): δ ϭ 0.88 (t, 3 H, aq. saturated NaHCO₃, and water, dried (MgSO₄), filtered, and con-

octyl CH₃), 1.08 (t, 3 H, COCH₂CH₃), 1.28 (m, 10 H, 5 octyl CH₂), centrated. Column chromatography (toluene/EtOAc, 3:1) of the resi-

1.59 (m, 2 H, octyl CH₂), 1.96, 1.99, 2.02, 2.04, 2.06, 2.07, 2.11, and due afforded 20, isolated as a syrup (233 mg, 74%). TLC (toluene/

2.14 (8 s, 3, 3, 3, 3, 3, 6, 3, 3 H, 9 COCH₃), 2.15 (m, 2 H, EtOAc, 3:1): R_fϭ 0.56. [α]²_D⁰ϭ Ϫ6 (c ϭ 1, CHCl₃). ¹H NMR

COCH₂CH₃), 3.40 (m, 1 H, octyl OCHH), 4.39 (dd, J_5Ј,6Јaϭ 2.5, (300 MHz, CDCl₃): δ ϭ 0.87 (t, 3 H, octyl CH₃), 1.06 (d, J_5ЈЈ,6ЈЈ

ϭ

J_6Јa,6Јbϭ 11.8 Hz, 1 H, 6Јa-H), 4.48 (d, J_1ЈЈ,2ЈЈϭ 7.7 Hz, 1 H, 1ЈЈ- 6.0 Hz, 3 H, 6ЈЈ-H), 1.23 (br. s, 10 H, 5 octyl CH₂), 1.42 (m, 2 H,

H), 4.69 (d, J_1,2ϭ 1.4 Hz, 1 H, 1-H), 4.73 (d, J_1Ј,2Јϭ 7.7 Hz, 1 H, octyl CH₂), 1.96, 2.01, and 2.07 (3 s, each 3 H, 3 COCH₃), 2.97 and

1Ј-H), 4.96 (dd, J_2ЈЈ,3ЈЈϭ 10.4, J_3ЈЈ,4ЈЈϭ 3.3 Hz, 1 H, 3ЈЈ-H), 5.15

3.19 (2 m, each 1 H, octyl OCH₂), 5.26 (d, J_1Ј,2Јϭ 7.4 Hz, 1 H, 1Ј-

(dd, 1 H, 4-H), 5.29 (dd, 1 H, 3Ј-H), 5.34 (d, J_4ЈЈ,5ЈЈϽ 1 Hz, 1 H, H), 6.80Ϫ7.42 (m, 25 H, 5 OCH₂C₆H₅), 7.45Ϫ7.70 (m, 4 H, Phth)

4ЈЈ-H), 5.69 (d, J_2Ј,NHϭ 8.5 Hz, 1 H, NH) ppm. ¹³C NMR ppm. MS (FAB^ϩ) of C₇₅H₈₇NO₁₉(1305): found m/z ϭ 1306 [M ϩ

(75.5 MHz, CDCl₃): δ ϭ 9.4 (COCH₂CH₃), 13.8 (octyl CH₃),

20.3Ϫ20.6 (COCH₃), 22.4, 25.9, 28.9, 29.0, 29.1, 29.4, and 31.6 (6

octyl CH₂, COCH₂CH₃), 54.0 (C-2Ј), 60.7, 62.6, 62.7, and 68.1 (C-

6, C-6Ј, C-6ЈЈ, octyl OCH₂), 66.0, 66.5, 68.3, 68.9, 70.0, 70.5, 70.6,

71.4, 72.5, 74.6, and 75.9 (C-2, C-3, C-4, C-5, C-3Ј, C-4Ј, C-5Ј, C-

2ЈЈ, C-3ЈЈ, C-4ЈЈ, C-5ЈЈ), 97.2, 99.2, and 100.7 (C-1, C-1Ј, C-1ЈЈ),

168.9, 169.3, 169.8, 169.9, 170.0 (2 C), 170.1, 170.2, and 170.5 (9

COCH₃), 173.8 (COCH₂CH₃) ppm. MS (FAB^ϩ) of C₄₇H₇₁NO₂₅

(1049): found m/z ϭ 1050 [M ϩ H]^ϩ, 1072 [M ϩ Na]^ϩ.

H]^ϩ, 1328 [M ϩ Na]^ϩ.

Octyl

(2,3,4-Tri-O-acetyl-β-

D

-fucopyranosyl)-(1Ǟ4)-(2-acetamido-

3,6-di-O-benzyl-2-deoxy-β-

D-glucopyranosyl)-(1Ǟ2)-3,4,6-tri-O-

benzyl-α-D-mannopyranoside (21): A solution of 20 (72 mg, 55 µmol)

˚

in 1-BuOH (8 mL), containing molecular sieves 3 A (0.25 g), was

stirred under Ar for 30 min, then 1,2-diaminoethane (0.74 mL,

11 mmol) was added. The mixture was stirred overnight at 90 °C,

filtered through Celite, and co-concentrated with toluene and EtOH.

A solution of the residue in pyridine/acetic anhydride (6 mL, 1:1)

Octyl β-

copyranosyl-(1Ǟ2)-α-

(88 mg, 84 µmol) in MeOH/CH₂Cl₂(10 mL, 4:1) was added NaOMe

D

-Galactopyranosyl-(1Ǟ4)-2-deoxy-2-propionamido-β-

D-glu- was stirred overnight, then co-concentrated with toluene. Column

D-mannopyranoside (2): To a solution of 17 chromatography (toluene/EtOAc, 3:1 Ǟ 1:1) of the residue yielded

21, isolated as a syrup (58 mg, 85%). TLC (toluene/EtOAc, 3:1):

(pH ϭ 9), and the mixture was stirred for 2.5 h. After neutralisation R_fϭ 0.29. [α]_D²⁰ϭ Ϫ3 (c ϭ 0.8, CHCl₃). ¹H NMR (300 MHz,

with Dowex 50 ϫ 8 (H^ϩ) and filtration, the solution was concen- CDCl₃): δ ϭ 0.88 (t, 3 H, octyl CH₃), 1.07 (d, J_5ЈЈ,6ЈЈϭ 6.0 Hz, 3 H,

trated. Gel-filtration of the residue on a Bio-Gel P-2 column, eluted 6ЈЈ-H), 1.26 (m, 10 H, 5 octyl CH₂), 1.50 (m, 2 H, octyl CH₂), 1.74

with H₂O, and subsequent lyophilization yielded 2, isolated as a

white powder (53 mg, 94%). TLC (CH₂Cl₂/MeOH/H₂O, 5:10:3):

R_fϭ 0.88. [α]_D²⁰ϭ Ϫ2 (c ϭ 1, H₂O). High-resolution MS of

(s, 3 H, NHCOCH₃), 1.96, 1.97, and 2.11 (3 s, each 3 H, 3 COCH₃),

4.13 (s, J_1,2Ͻ 1 Hz, 1 H, 1-H), 5.73 (d, J_2Ј,NHϭ 8.7 Hz, 1 H, NH),

7.10Ϫ7.58 (m, 25 H,

5

OCH₂C₆H₅) ppm. MS (FAB^ϩ) of

C₂₉H₅₃NO₁₆(671.3364): calcd. 694.3262, found 694.3265 [M ϩ Na]. C₆₉H₈₇NO₁₈(1217): found m/z ϭ 1218 [M ϩ H]^ϩ, 1240 [M ϩ Na]^ϩ.

For ¹H and ¹³C NMR spectroscopic data, see Tables 1 and 2, respec-

Octyl

3,6-di-O-acetyl-2-deoxy-β-

-fucopyranoside (19): A solution acetyl-α- -mannopyranoside (22): To a solution of 21 (106 mg, 86

(2,3,4-Tri-O-acetyl-β-D-fucopyranosyl)-(1Ǟ4)-(2-acetamido-

tively.

D

-glucopyranosyl)-(1Ǟ2)-3,4,6-tri-O-

Ethyl 2,3,4-Tri-O-acetyl-1-thio-β-

D

of -fucose (18; 936 mg, 5.7 mmol) in pyridine/acetic anhydride

(20 mL, 1:1) was stirred for 7 h, then co-concentrated with toluene.

The residue was dissolved in dry CH₂Cl₂(10 mL), containing molec-

µmol) in EtOH/EtOAc (15 mL, 1:1) were added 10% Pd/C (48 mg)

and HOAc (165 µL), and the mixture was stirred overnight under

H₂, then filtered through Celite, and concentrated. A solution of the

residue in pyridine/acetic anhydride (43 mL, 1:1) was stirred over-

night, then co-concentrated with toluene. Column chromatography

(toluene/EtOAc, 1:3) of the residue yielded 22, isolated as a syrup

(61 mg, 72%). TLC (toluene/EtOAc, 1:3): R_fϭ 0.17. [α]_D²⁰ϭ Ϫ21

(c ϭ 1, CHCl₃). ¹H NMR (300 MHz, CDCl₃): δ ϭ 0.88 (t, 3 H,

˚

ular sieves 4 A (200 mg), and ethanethiol (507 µL, 6.84 mmol) was

added. The mixture was stirred under Ar for 60 min, then cooled to

0 °C, and BF₃·Et₂O (2.86 mL, 22.8 mmol) was added. After 100 min,

during which period the temperature was allowed to reach room

temperature, the mixture was neutralised with NEt₃, and filtered.

The filtrate was diluted with CH₂Cl₂, washed with water and aq. octyl CH₃), 1.19 (d, J_5ЈЈ,6ЈЈϭ 6.4 Hz, 3 H, 6ЈЈ-H), 1.30 (m, 10 H, 5

saturated NaHCO₃, dried (MgSO₄), and concentrated. Column octyl CH₂), 1.58 (m, 2 H, octyl CH₂), 1.95, 1.98, 2.01, 2.03, 2.08,

chromatography (toluene/EtOAc, 4:1) of the residue yielded 19, iso- 2.10, and 2.15 (7 s, 3, 3, 3, 6, 6, 3, 3 H, 9 COCH₃), 3.41 (m, 1 H,

lated as a white foam (1.16 g, 61%), and the α-product (700 mg, octyl OCHH), 4.44 (d, J_1ЈЈ,2ЈЈϭ 7.8 Hz, 1 H, 1ЈЈ-H), 4.63 (d, J_1Ј,2Ј

ϭ

36%). TLC (toluene/EtOAc, 4:1): R_fϭ 0.42. [α]²_D⁰ϭ Ϫ28 (c ϭ 1,

7.4 Hz, 1 H, 1Ј-H), 4.71 (s, J_1,2Ͻ 1 Hz, 1 H, 1-H), 5.68 (d, J_2Ј,NHϭ

1

CHCl₃). H NMR (300 MHz, CDCl₃): δ ϭ 0.94 (d, J_5,6ϭ 6.3 Hz,

8.7 Hz, 1 H, NH) ppm. MS (FAB^ϩ) of C₄₄H₆₇NO₂₃(977): found

3 H, 6-H), 1.02 (t, 3 H, SCH₂CH₃), 1.70, 1.79, and 1.89 (3 s, each 3 m/z ϭ 978 [M ϩ H]^ϩ, 1000 [M ϩ Na]^ϩ.

H, 3 COCH₃), 2.37Ϫ2.57 (m, 2 H, SCH₂CH₃), 3.63 (m, 1 H, 5-H),

Octyl β-D-Fucopyranosyl-(1Ǟ4)-2-acetamido-2-deoxy-β-

D-glucopyr-

4.26 (d, J_1,2ϭ 9.7 Hz, 1 H, 1-H), 4.83 (dd, J_2,3ϭ 10.0, J_3,4ϭ 3.2 Hz,

1 H, 3-H), 4.95 (t, 1 H, 2-H), 5.01 (dd, J_4,5Ͻ 1 Hz, 1 H, 4-H) ppm.

MS (FAB^ϩ) of C₁₄H₂₂O₇S (334): found m/z ϭ 335 [M ϩ H]^ϩ, 357

[M ϩ Na]^ϩ.

anosyl-(1Ǟ2)-α-D-mannopyranoside (3): To a solution of 22 (61 mg,

62 µmol) in MeOH/CH₂Cl₂(8 mL, 3:1) was added NaOMe (pH ϭ

9), and the mixture was stirred for 5 h. After neutralisation with

Dowex 50 ϫ 8 (H^ϩ) and filtration, the solution was concentrated.

Octyl

benzyl-2-deoxy-2-phthalimido-β-

O-benzyl-α- -mannopyranoside (20): A solution of 19 (109 mg, 325 powder (24 mg, 61%). TLC (1-BuOH/EtOH/HOAc/H₂O, 4:2:2:1):

(2,3,4-Tri-O-acetyl-β-D-fucopyranosyl)-(1Ǟ4)-(3,6-di-O- Gel-filtration of the residue on a Bio-Gel P-2 column, eluted with

D

-glucopyranosyl)-(1Ǟ2)-3,4,6-tri- water, and subsequent lyophilization yielded 3, isolated as a white

D

Eur. J. Org. Chem. 2003, 3569Ϫ3586

www.eurjoc.org

 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

3579

J. P. Kamerling et al.

FULL PAPER

R_fϭ 0.32. [α]_D²⁰ϭ Ϫ7 (c ϭ 1, H₂O). High-resolution MS of

and 1.52 (4 s, each 3 H, 4 COCH₃), 3.33 (dd, J_5,6aϭ 5.4, J_6a,6b

ϭ

C₂₈H₅₁NO₁₅(641.3259): calcd. 664.3156, found 664.3170 [M ϩ Na].

12.7 Hz, 1 H, 6a-H), 3.47 (dd, J_5,6bϭ 7.8 Hz, 1 H, 6b-H), 3.89 (ddd,

For ¹H and ¹³C NMR spectroscopic data, see Tables 1 and 2, respec- 1 H, 5-H), 4.17 (dd, J_3,4ϭ 7.9, J_4,5ϭ 2.0 Hz, 1 H, 4-H), 4.30 (dd,

tively.

J_1,2ϭ 5.0, J_2,3ϭ 2.5 Hz, 1 H, 2-H), 4.60 (dd, 1 H, 3-H), 5.52 (d, 1

H, 1-H) ppm. MS (FAB^ϩ) of C₁₂H₁₉N₃O₅(285): found m/z ϭ 286

[M ϩ H]^ϩ, 308 [M ϩ Na]^ϩ.

Octyl (2,3,4-Tri-O-acetyl-β-

tyl-2-deoxy-2-propionamido-β-

D

-fucopyranosyl)-(1Ǟ4)-(3,6-di-O-ace-

D-glucopyranosyl)-(1Ǟ2)-3,4,6-tri-O-

acetyl-α-

D

-mannopyranoside (24): A solution of 20 (161 mg, 123 2,3,4-Tri-O-acetyl-6-azido-6-deoxy-α-

D

-galactopyranosyl

Trichlo-

˚

µmol) in 1-BuOH (17 mL), containing molecular sieves 3 A (0.55 g),

was stirred for 30 min under Ar, then 1,2-diaminoethane (1.66 mL,

25 mmol) was added. The mixture was stirred at 90 °C overnight,

filtered through Celite, and co-concentrated with toluene and EtOH.

A solution of the residue in pyridine/propionic anhydride (12 mL,

1:1) was stirred overnight, then co-concentrated with toluene and

EtOH. Column chromatography (toluene/EtOAc, 3:1) of the residue

roacetimidate (30): To a solution of 27 (900 mg, 3.15 mmol) in

CH₂Cl₂(30 mL) were added trifluoroacetic acid (5.6 mL, 73 mmol)

and water (0.7 mL), and the mixture was stirred for 2 h, then co-

concentrated with toluene. A solution of the residue in pyridine/

acetic anhydride (20 mL, 1:1) was stirred overnight, then co-concen-

trated with toluene. Column chromatography (toluene/EtOAc, 3:1)

of the residue yielded 28, isolated as a syrup (583 mg, 50%). To a

yielded 23, isolated as a syrup (138 mg, 87%). To a solution of 23 in solution of 28 in DMF (3 mL) was added hydrazinium acetate

EtOH/EtOAc (16 mL, 1:1) were added 10% Pd/C (60 mg) and

HOAc (0.2 mL), and the mixture was stirred overnight under H₂,

then filtered through Celite, and concentrated. To a solution of the

(158 mg, 1.72 mmol), and the solution was stirred for 70 min at 50

°C, then diluted with EtOAc, washed with water and aq. saturated

NaCl, dried (MgSO₄), and concentrated to give 29, isolated as a

residue in MeOH (15 mL) was added NaOMe (pH ϭ 9), and the foam (461 mg, 89%). To a solution of 29 (461 mg, 1.4 mmol) in

mixture was stirred for 6 h, then neutralised with Dowex 50 ϫ 8 CH₂Cl₂(8 mL) was added, at 0 °C, trichloroacetonitrile (1.4 mL,

(H^ϩ), and concentrated. The residue was dissolved in pyridine/acetic

anhydride (16 mL, 1:1) and stirred overnight, then co-concentrated

with toluene. Column chromatography (toluene/EtOAc, 1:2) of the

residue yielded 24, isolated as a syrup (58 mg, 55%). TLC (toluene/

EtOAc, 1:2): R_fϭ 0.25. [α]²_D⁰ϭ Ϫ18 (c ϭ 1, CHCl₃). ¹H NMR

14 mmol) and 1,8-diazabicyclo[5.4.0]undec-7-ene (104 µL,

0.7 mmol), and the mixture was stirred for 1 h, then concentrated.

Column chromatography (toluene/EtOAc, 4:1) of the residue yielded

30, isolated as a syrup (380 mg, 63%). TLC (toluene/EtOAc, 4:1):

R_fϭ 0.33. [α]_D²⁰ϭ ϩ68 (c ϭ 1, CHCl₃). ¹H NMR (300 MHz,

(300 MHz, CDCl₃): δ ϭ 0.89 (t, 3 H, octyl CH₃), 1.11 (t, 3 H, CDCl₃): δ ϭ 1.94, 1.97, and 2.12 (3 s, each 3 H, 3 COCH₃), 3.16

COCH₂CH₃), 1.21 (d, J_5ЈЈ,6ЈЈϭ 5.8 Hz, 3 H, 6ЈЈ-H), 1.33 (m, 10 H,

5 octyl CH₂), 1.59 (m, 2 H, octyl CH₂), 1.89Ϫ2.16 (m, 26 H, 8 7.7 Hz, 1 H, 6b-H), 4.30 (m, 1 H, 5-H), 5.28 (dd, J_1,2ϭ 3.3, J_2,3

COCH₃, COCH₂CH₃), 3.39 (m, 1 H, octyl OCHH), 4.42 (d, J_1ЈЈ,2ЈЈ

(dd, J_5,6aϭ 4.9, J_6a,6bϭ 12.9 Hz, 1 H, 6a-H), 3.39 (dd, J_5,6b

ϭ

10.8 Hz, 1 H, 2-H), 5.35 (dd, J_3,4ϭ 2.9 Hz, 1 H, 3-H), 5.47 (dd,

7.8 Hz, 1 H, 1ЈЈ-H), 4.62 (d, J_1Ј,2Јϭ 7.3 Hz, 1 H, 1Ј-H), 4.67 (s, J_1,2

J_4,5ϭ 1.1 Hz, 1 H, 4-H), 6.55 (d, 1 H, 1-H), 8.60 (s, 1 H, NH) ppm.

Ͻ 1 Hz, 1 H, 1-H), 5.60 (d, J_2Ј,NHϭ 7.4 Hz, 1 H, NH) ppm. MS MS (FAB^ϩ) of C₁₄H₁₇Cl₃N₄O₈(474): found m/z ϭ 475 [M ϩ H]^ϩ,

(FAB^ϩ) of C₄₅H₆₉NO₂₃(991): found m/z ϭ 992 [M ϩ H]^ϩ, 1014 [M 497 [M ϩ Na]^ϩ.

ϩ Na]^ϩ.

Octyl

-gluco- (1Ǟ4)-(3,6-di-O-benzyl-2-deoxy-2-phthalimido-β-

-mannopyranoside (4): To a solution of 24 (1Ǟ2)-3,4,6-tri-O-benzyl-α- -mannopyranoside (31): A solution of 13

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

D-galactopyranosyl)-

Octyl β-

pyranosyl-(1Ǟ2)-α-

D

-Fucopyranosyl-(1Ǟ4)-2-deoxy-2-propionamido-β-

D

D-glucopyranosyl)-

D

(58 mg, 58 µmol) in MeOH (10 mL) was added NaOMe (pH ϭ 9), (260 mg, 251 µmol) and 30 (195 mg, 450 µmol) in CH₂Cl₂(12.5 mL),

˚

and the mixture was stirred for 24 h. After neutralisation with

containing molecular sieves 4 A (50 mg), was stirred under Ar at 0

Dowex 50 ϫ 8 (H^ϩ) and filtration, the solution was concentrated.

°C for 2.5 h, then TMSOTf (17 µL, 80 µmol) was added. After stir-

Gel-filtration of the residue on a Bio-Gel P-2 column, eluted with ring for 35 min, the mixture was neutralised with NEt₃, filtered, di-

water, and subsequent lyophilization yielded 4, isolated as a white

powder (36 mg, 93%). TLC (1-BuOH/EtOH/HOAc/H₂O, 4:2:2:1):

R_fϭ 0.38. [α]_D²⁰ϭ Ϫ1 (c ϭ 1, H₂O). High-resolution MS of

C₂₉H₅₃NO₁₅(655.3415): calcd. 678.3313, found 678.3331 [M ϩ Na].

luted with CH₂Cl₂, washed with water, dried (MgSO₄), and concen-

trated. Column chromatography (toluene/EtOAc, 3:1) of the residue

yielded 31, isolated as a syrup (206 mg, 61%). TLC (toluene/EtOAc,

1

3:1): R_fϭ 0.59. [α]_D²⁰ϭ ϩ1 (c ϭ 1, CHCl₃). H NMR (300 MHz,

For ¹H and ¹³C NMR spectroscopic data, see Tables 1 and 2, respec- CDCl₃): δ ϭ 0.90 (t, 3 H, octyl CH₃), 1.25 (m, 10 H, 5 octyl CH₂),

tively.

1.46 (m, 2 H, octyl CH₂), 2.00, 2.05, and 2.10 (3 s, each 3 H, 3

COCH₃), 4.67 (d, J_1ЈЈ,2ЈЈϭ 8.0 Hz, 1 H, 1ЈЈ-H), 5.17 (dd, J_2ЈЈ,3ЈЈ

10.4 Hz, 1 H, 2ЈЈ-H), 5.28 (d, J_1Ј,2Јϭ 7.6 Hz, 1 H, 1Ј-H), 5.28 (d,

J_3ЈЈ,4ЈЈϭ 3.0, J_4ЈЈ,5ЈЈϽ 1 Hz, 1 H, 4ЈЈ-H), 6.90Ϫ7.48 (m, 25 H, 5

OCH₂C₆H₅), 7.51Ϫ7.68 (2 m, each 2 H, Phth) ppm. MS (FAB^ϩ) of

C₇₅H₈₆N₄O₁₉(1346): found m/z ϭ 1347 [M ϩ H]^ϩ, 1369 [M ϩ Na]^ϩ.

ϭ

6-Azido-6-deoxy-1,2:3,4-di-O-isopropylidene-α-D-galactopyranose

(27): To a solution of 1,2:3,4-di-O-isopropylidene-α--galactopyr-

anose (25; 1.08 g, 4.15 mmol) in pyridine (10 mL) was added drop-

wise a solution of p-toluenesulfonyl chloride (1.3 g, 6.3 mmol) in

CH₂Cl₂(10 mL), and the mixture was stirred for 5 h. After the ad-

dition of water (1 mL), the mixture was stirred for 10 min, then co-

concentrated with toluene. A solution of the residue in CH₂Cl₂was

Octyl

(1Ǟ4)-(2-acetamido-3,6-di-O-benzyl-2-deoxy-β-

-mannopyranoside (32): A solution of 31

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

D-galactopyranosyl)-

D-glucopyranosyl)-

washed with water and aq. NaHCO₃, dried (MgSO₄), and concen- (1Ǟ2)-3,4,6-tri-O-benzyl-α-

D

trated. Column chromatography (toluene/EtOAc, 5:1) of the residue

yielded 26, isolated as a syrup (1.59 g, 92%). To a solution of 26 3 A (0.3 g), was stirred under Ar for 30 min, then 1,2-diaminoethane

(103 mg, 76 µmol) in 1-BuOH (10 mL), containing molecular sieves

˚

(1.31 g, 3.17 mmol) in Me₂SO (30 mL) was added NaN₃(1 g,

15.7 mmol), and the mixture was stirred for 2 h at 160 °C, then

cooled to room temperature, poured into iced water, extracted with

EtOAc, and the organic phase was dried (MgSO₄), and concentrated

(1.05 mL, 15.7 mmol) was added. The mixture was stirred overnight

at 90 °C, filtered through Celite, and co-concentrated with toluene

and EtOH. A solution of the residue in pyridine/acetic anhydride

(30 mL, 1:1) was stirred overnight, then co-concentrated with tolu-

to yield 27, isolated as a syrup (900 mg, 99%). TLC (toluene/EtOAc, ene and EtOH. Column chromatography (toluene/EtOAc, 2:1) of the

3:1): R_fϭ 0.62. IR (KBr): ν˜ ϭ 2110 cm^Ϫ1(N₃). [α]²_D⁰ϭ ϩ43 (c ϭ

residue yielded 32, isolated as a syrup (96 mg, 99%). TLC (toluene/

1, CHCl₃). H NMR (300 MHz, [D₆]Me₂SO): δ ϭ 1.31, 1.32, 1.43, EtOAc, 2:1): R_fϭ 0.50. [α]²_D⁰ϭ ϩ5 (c ϭ 1, CHCl₃). ¹H NMR

1

3580

 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.eurjoc.org

Eur. J. Org. Chem. 2003, 3569Ϫ3586

β-D-Galp-(1Ǟ4)-β-D-GlcpNAc-(1Ǟ2)-α-D-Manp-(1ǞO)(CH₂)₇CH₃Mimics

(300 MHz, CDCl₃): δ ϭ 0.88 (t, 3 H, octyl CH₃), 1.27 (m, 10 H, 5 solution of 34 (91 mg, 71 µmol) in MeOH (15 mL) was added Na-

FULL PAPER

octyl CH₂), 1.51 (m, 2 H, octyl CH₂), 1.72 (s, 3 H, NHCOCH₃),

OMe (pH ϭ 9), and the mixture was stirred for 2.5 h. After neutralis-

1.96, 2.01, and 2.10 (3 s, each 3 H, 3 COCH₃), 4.11 (d, J_1,2Ͻ 1 Hz, ation with Dowex 50 ϫ 8 (H^ϩ) and filtration, the solution was con-

1 H, 1-H), 5.23 (d, J_3ЈЈ,4ЈЈϭ 3.3, J_4ЈЈ,5ЈЈϽ 1 Hz, 1 H, 4ЈЈ-H), 5.69 (d, centrated. Column chromatography (CH₂Cl₂/MeOH, 9:1) of the

J_2Ј,NHϭ 7.1 Hz, 1 H, NH), 7.15Ϫ7.45 (m, 25 H, 5 OCH₂C₆H₅)

ppm. MS (FAB^ϩ) of C₆₉H₈₆N₄O₁₈(1258): found m/z ϭ 1259 [M ϩ

H]^ϩ, 1281 [M ϩ Na]^ϩ.

residue yielded 35, isolated as a syrup (65 mg, 79%). To a solution

of 35 (65 mg, 53 µmol) in 2-propanol/water (10 mL, 3:2) were added

10% Pd/C (160 mg) and aq. 25% NH₃(pH ϭ 9), and the mixture

was stirred under H₂for 3.5 h. After the removal of NH₃by bubbling

N₂through the mixture, the solution was acidified with HOAc,

stirred under H₂for 23 h, filtered through Celite, and concentrated.

Gel-filtration of the residue on a Bio-Gel P-2 column, eluted with

water, yielded crude 6. The residue was dissolved in water (1 mL),

and applied on a SepPak C₁₈cartridge that was eluted first with

water, then with MeOH. The MeOH fraction was concentrated and

lyophilized to yield 6, isolated as a white powder (18 mg, 48%). TLC

(1-BuOH/EtOH/HOAc/H₂O, 4:2:2:1): R_fϭ 0.15. [α]²_D⁰ϭ Ϫ1 (c ϭ 1,

H₂O). High-resolution MS of C₂₉H₅₄N₂O₁₅(670.3524): calcd.

Octyl (6-Azido-6-deoxy-β-

3,6-di-O-benzyl-2-deoxy-β-

benzyl-α- -mannopyranoside (33): To a solution of 32 (96 mg, 76

D-galactopyranosyl)-(1Ǟ4)-(2-acetamido-

D

-glucopyranosyl)-(1Ǟ2)-3,4,6-tri-O-

D

µmol) in MeOH (15 mL) was added NaOMe (pH ϭ 9), and the

mixture was stirred for 2 h. After neutralisation with Dowex 50 ϫ 8

(H^ϩ), and filtration, the solution was concentrated. Column chroma-

tography (CH₂Cl₂/MeOH, 95:5) of the residue yielded 33, isolated

as a syrup (61 mg, 70%). TLC (CH₂Cl₂/MeOH, 95:5): R_fϭ 0.77.

1

[α]²_D⁰ϭ Ϫ8 (c ϭ 1, MeOH). H NMR (300 MHz, CDCl₃): δ ϭ 0.90

(t, 3 H, octyl CH₃), 1.27 (m, 10 H, 5 octyl CH₂), 1.52 (m, 2 H, octyl

CH₂), 1.66 (s, 3 H, NHCOCH₃), 3.04 (m, 1 H, octyl OCHH), 4.13

(d, J_1,2Ͻ 1 Hz, 1 H, 1-H), 5.13 (d, J_1Ј,2Јϭ 7.3 Hz, 1 H, 1Ј-H), 5.87

(d, J_2Ј,NHϭ 7.1 Hz, 1 H, NH), 7.25Ϫ7.40 (m, 25 H, 5 OCH₂C₆H₅)

ppm. MS (FAB^ϩ) of C₆₃H₈₀N₄O₁₅(1132): found m/z ϭ 1133 [M ϩ

H]^ϩ, 1155 [M ϩ Na]^ϩ.

1

671.3602, found 671.3597 [M ϩ H]. For H and ¹³C NMR spectro-

scopic data, see Tables 1 and 2, respectively.

Octyl (2-O-Acetyl-3,4,6-tri-O-benzyl-β-

(3,6-di-O-benzyl-2-deoxy-2-phthalimido-β-

3,4,6-tri-O-benzyl-α- -mannopyranoside (37): A solution of ethyl 2-

O-acetyl-3,4,6-tri-O-benzyl-1-thio-β--galactopyranoside

36^[17]

D-galactopyranosyl)-(1Ǟ4)-

D

-glucopyranosyl)-(1Ǟ2)-

D

Octyl 6-Amino-6-deoxy-β-D-galactopyranosyl-(1Ǟ4)-2-acetamido-2-

(486 mg, 906 µmol) and 13 (541 mg, 522 µmol) in CH₂Cl₂(25 mL),

deoxy-β- -glucopyranosyl-(1Ǟ2)-α-

D

D-mannopyranoside (5): To a

˚

containing powdered molecular sieves 4 A (0.5 g), was stirred under

solution of 33 (61 mg, 53 µmol) in 2-propanol (6 mL) and water

(4 mL) were added 10% Pd/C (160 mg) and aq. 25% NH₃(pH ϭ

10), and the mixture was stirred under H₂for 3 h. After the removal

of NH₃by bubbling N₂through the mixture, the solution was acidi-

fied with HOAc, stirred under H₂for 23 h, filtered through Celite,

and concentrated. Gel-filtration of the residue on a Bio-Gel P-2 col-

umn, eluted with water, yielded crude 5. The residue was dissolved

in water (1 mL), and applied on a SepPak C₁₈cartridge that was

eluted first with water, then with MeOH. The MeOH fraction was

concentrated and lyophilized to yield 5, isolated as a white powder

(34.5 mg, 99%). TLC (1-BuOH/EtOH/HOAc/H₂O, 4:2:2:1): R_fϭ

0.16. [α]²_D⁰ϭ Ϫ15 (c ϭ 1, H₂O). High-resolution MS of C₂₈H₅₂N₂O₁₅

Ar for 3 h. The mixture was cooled to 0 °C, then N-iodosuccinimide

(289 mg, 1.28 mmol) and triflic acid (16 µL, 181 µmol) were added.

After 40 min, the solution was neutralised with NEt₃, filtered, di-

luted with CH₂Cl₂, washed with aq. 10% NaHSO₃, aq. saturated

NaHCO₃, and water, dried (MgSO₄), and concentrated. Column

chromatography (toluene/EtOAc, 8:1) of the residue yielded 37, iso-

lated as a syrup (548 mg, 70%). TLC (toluene/EtOAc, 6:1): R_fϭ

0.61. [α]²_D⁰ϭ ϩ4 (c ϭ 1, CHCl₃). ¹H NMR (300 MHz, CDCl₃): δ ϭ

0.88 (t, 3 H, octyl CH₃), 1.22 (m, 10 H, 5 octyl CH₂), 1.41 (m, 2 H,

octyl CH₂), 1.98 (s, 3 H, COCH₃), 2.75 and 3.19 (2 m, each 1 H,

octyl OCH₂), 5.23 (d, J_1Ј,2Јϭ 8.0 Hz, 1 H, 1Ј-H), 5.38 (dd, J_1ЈЈ,2ЈЈ

ϭ

7.7, J_2ЈЈ,3ЈЈϭ 9.9 Hz, 1 H, 2ЈЈ-H), 6.75Ϫ7.38 (m, 40 H, 8

OCH₂C₆H₅), 7.45Ϫ7.70 (m, 4 H, Phth) ppm. MS (FAB^ϩ) of

C₉₂H₁₀₁NO₁₈(1507): found m/z ϭ 1508 [M ϩ H]^ϩ, 1530 [M ϩ Na]^ϩ.

1

(656.3368): calcd. 657.3446, found 657.3441 [M ϩ H]. For H and

¹³C NMR spectroscopic data, see Tables 1 and 2, respectively.

Octyl (6-Azido-6-deoxy-2,3,4-tri-O-propionyl-β-

(1Ǟ4)-(3,6-di-O-benzyl-2-deoxy-2-propionamido-β-

ranosyl)-(1Ǟ2)-3,4,6-tri-O-benzyl-α- -mannopyranoside (34): A solu-

D-galactopyranosyl)-

Octyl (2-O-Acetyl-3,4,6-tri-O-benzyl-β-

acetamido-3,6-di-O-benzyl-2-deoxy-β-

3,4,6-tri-O-benzyl-α- -mannopyranoside (38): A solution of 37

D-galactopyranosyl)-(1Ǟ4)-(2-

D

-glucopy-

D

-glucopyranosyl)-(1Ǟ2)-

D

tion of 31 (103 mg, 76 µmol) in 1-BuOH (10 mL), containing molec-

˚

(238 mg, 158 µmol) in 1-BuOH (22 mL), containing molecular

ular sieves 3 A (0.3 g), was stirred under Ar for 30 min, then 1,2-

˚

sieves 3 A (0.2 g), was stirred under Ar for 1 h, then 1,2-diamino-

diaminoethane (1.05 mL, 15.7 mmol) was added. The mixture was

stirred at 90 °C overnight, filtered through Celite, and co-concen-

trated with toluene and EtOH. A solution of the residue in pyridine/

propionic anhydride (30 mL, 1:1) was stirred overnight, then co-con-

centrated with toluene and EtOH. Column chromatography (tolu-

ene/EtOAc, 4:1) of the residue yielded 34, isolated as a syrup (91 mg,

94%). TLC (toluene/EtOAc, 3:1): R_fϭ 0.56. [α]²_D⁰ϭ ϩ7 (c ϭ 1,

CHCl₃). ¹H NMR (300 MHz, CDCl₃): δ ϭ 0.88 (t, 3 H, octyl CH₃),

0.94Ϫ1.23 (m, 12 H, 3 COCH₂CH₃, NHCOCH₂CH₃), 1.29 (m, 10

H, 5 octyl CH₂), 1.51 (m, 2 H, octyl CH₂), 1.95 (m, 2 H,

NHCOCH₂CH₃), 2.18Ϫ2.53 (m, 6 H, 3 COCH₂CH₃), 4.12 (d, J_1,2

Ͻ 1 Hz, 1 H, 1-H), 4.87 (dd, J_2ЈЈ,3ЈЈϭ 10.3, J_3ЈЈ,4ЈЈϭ 3.1 Hz, 1 H,

3ЈЈ-H), 5.14 (dd, J_1ЈЈ,2ЈЈϭ 8.0 Hz, 1 H, 2ЈЈ-H), 5.18 (d, J_1Ј,2Јϭ 7.7 Hz,

ethane (2.0 mL, 30 mmol) was added. The mixture was stirred over-

night at 90 °C, filtered through Celite, and co-concentrated with

toluene. A solution of the residue in pyridine/acetic anhydride

(60 mL, 1:1) was stirred overnight, then co-concentrated with tolu-

ene to give 38, isolated as a syrup (211 mg, 94%). TLC (toluene/

EtOAc, 3:1): R_fϭ 0.29. [α]²_D⁰ϭ Ϫ1 (c ϭ 1, CHCl₃). ¹H NMR

(300 MHz, CDCl₃): δ ϭ 0.87 (t, 3 H, octyl CH₃), 1.25 (m, 10 H, 5

octyl CH₂), 1.52 (m, 2 H, octyl CH₂), 1.76 (s, 3 H, NHCOCH₃),

1.96 (s, 3 H, COCH₃), 4.11 (s, J_1,2Ͻ 1 Hz, 1 H, 1-H), 5.33 (dd,

J_1ЈЈ,2ЈЈϭ 8.1, J_2ЈЈ,3ЈЈϭ 10.1 Hz, 1 H, 2ЈЈ-H), 5.82 (d, J_2Ј,NH

ϭ

7.8 Hz, 1 H, NH), 7.10Ϫ7.45 (m, 40 H, 8 OCH₂C₆H₅) ppm. MS

(FAB^ϩ) of C₈₆H₁₀₁NO₁₇(1419): found m/z ϭ 1420 [M ϩ H]^ϩ, 1442

[M ϩ Na]^ϩ.

1 H, 1Ј-H), 5.24 (d, J_4ЈЈ,5ЈЈϽ 1 Hz, 1 H, 4ЈЈ-H), 5.74 (d, J_2Ј,NH

ϭ

6.8 Hz, 1 H, NH), 7.13Ϫ7.42 (m, 25 H, 5 OCH₂C₆H₅) ppm. MS

(FAB^ϩ) of C₇₃H₉₄N₄O₁₈(1314): found m/z ϭ 1315 [M ϩ H]^ϩ, 1337

[M ϩ Na]^ϩ.

Octyl (3,4,6-Tri-O-benzyl-2-O-phenoxythiocarbonyl-β-

anosyl)-(1Ǟ4)-(2-acetamido-3,6-di-O-benzyl-2-deoxy-β-

anosyl)-(1Ǟ2)-3,4,6-tri-O-benzyl-α- -mannopyranoside (40): To a

D

-galactopyr-

D

-glucopyr-

D

Octyl 6-Amino-6-deoxy-β-

D

-galactopyranosyl-(1Ǟ4)-2-deoxy-2-pro-

solution of 38 (211 mg, 148 µmol) in MeOH/CH₂Cl₂(100 mL, 3:1)

was added NaOMe (pH ϭ 10), and the mixture was stirred at 40

pionamido-β- -glucopyranosyl-(1Ǟ2)-α-

D

D-mannopyranoside (6): To a

Eur. J. Org. Chem. 2003, 3569Ϫ3586

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 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

3581

J. P. Kamerling et al.

FULL PAPER

°C for 17 h, then neutralised with Dowex 50 ϫ 8 (H^ϩ), filtered, and

concentrated to give 39. To a solution of the residue in acetonitrile

neutralised with Dowex 50 ϫ 8 (H^ϩ), filtered, and concentrated.

To a solution of the residue (43) in dry acetonitrile (22 mL) were

(25 mL) were added 4-(dimethylamino)pyridine (825 mg, added 4-(dimethylamino)pyridine (825 mg, 6.75 mmol) and phenyl

6.75 mmol) and phenyl chlorothionocarbonate (540 µL,

3.90 mmol), and the mixture was stirred at 100 °C for 4.5 h, then

chlorothionocarbonate (534 µL, 3.86 mmol), and the mixture was

stirred at 100 °C for 2 h, then overnight at room temperature. After

at room temperature overnight. After dilution with CH₂Cl₂, the dilution with CH₂Cl₂, the solution was washed with aq. 0.5  HCl,

solution was washed with aq. 0.5  HCl, aq. saturated NaHCO₃, aq. saturated NaHCO₃, and water, dried (MgSO₄), and concen-

and water, dried (MgSO₄), and concentrated. Column chromatog-

trated. Column chromatography (toluene/EtOAc, 5:1) of the resi-

due yielded 44, isolated as a syrup (128 mg, 43%). TLC (toluene/

EtOAc, 5:1): R_fϭ 0.61. [α]²_D⁰ϭ Ϫ2 (c ϭ 1, CHCl₃). ¹H NMR

(300 MHz, CDCl₃): δ ϭ 0.88 (t, 3 H, octyl CH₃), 0.97 (t, 3 H,

raphy (toluene/EtOAc, 4:1) of the residue yielded 40, isolated as a

syrup (139 mg, 61%). TLC (toluene/EtOAc, 4:1): R_fϭ 0.50. [α]²_D⁰

Ϫ7 (c ϭ 1, CHCl₃). H NMR (300 MHz, CDCl₃): δ ϭ 0.91 (t, 3

ϭ

1

H, octyl CH₃), 1.31 (m, 10 H, 5 octyl CH₂), 1.58 (m, 2 H, octyl NHCOCH₂CH₃), 1.26 (m, 10 H, 5 octyl CH₂), 1.52 (m, 2 H, octyl

CH₂), 1.73 (s, 3 H, NHCOCH₃), 3.22 (m, 1 H, octyl OCHH), 4.13 CH₂), 1.82Ϫ2.05 (m, 2 H, NHCOCH₂CH₃), 3.17 (m, 1 H, octyl

(s, J_1,2Ͻ 1 Hz, 1 H, 1-H), 5.18 (d, J_1Ј,2Јϭ 7.1 Hz, 1 H, 1Ј-H), 5.66 OCHH), 4.13 (s, J_1,2Ͻ 1 Hz, 1 H, 1-H), 5.18 (d, J_1Ј,2Јϭ 7.9 Hz, 1

(d, J_2Ј,NHϭ 7.1 Hz, 1 H, NH), 5.95 (dd, J_1ЈЈ,2ЈЈϭ 7.8, J_2ЈЈ,3ЈЈ

ϭ

H, 1Ј-H), 5.62 (d, J_2Ј,NHϭ 6.8 Hz, 1 H, NH), 5.90 (dd, J_1ЈЈ,2ЈЈϭ

10.0 Hz, 1 H, 2ЈЈ-H), 7.00Ϫ7.50 (m, 45 H, 9 OCH₂C₆H₅) ppm. MS 7.8, J_2ЈЈ,3ЈЈϭ 9.9 Hz, 1 H, 2ЈЈ-H), 6.92Ϫ7.47 (m, 45 H, 9

(FAB^ϩ) of C₉₁H₁₀₃NO₁₇S (1513): found m/z ϭ 1514 [M ϩ H]^ϩ,

1536 [M ϩ Na]^ϩ.

OCH₂C₆H₅) ppm. MS (FAB^ϩ) of C₉₂H₁₀₅NO₁₇S (1527): found

m/z ϭ 1528 [M ϩ H]^ϩ, 1550 [M ϩ Na]^ϩ.

Octyl (2-Deoxy-3,4,6-tri-O-benzyl-β-

(2-acetamido-3,6-di-O-benzyl-2-deoxy-β-

3,4,6-tri-O-benzyl-α- -mannopyranoside (41): To a solution of 40

(139 mg, 91 µmol) in dry toluene (3 mL) was added tributylstan-

nane (295 µL, 1.1 mmol), and the mixture was stirred at 100 °C. stannane (270 µL, 1.0 mmol), and the mixture was stirred at 100

D

-lyxo-hexopyranosyl)-(1Ǟ4)- Octyl (2-Deoxy-3,4,6-tri-O-benzyl-β-

-glucopyranosyl)-(1Ǟ2)- (3,6-di-O-benzyl-2-deoxy-2-propionamido-β-

(1Ǟ2)-3,4,6-tri-O-benzyl-α- -mannopyranoside (45): To a solution

of 44 (127 mg, 83 µmol) in dry toluene (3 mL) was added tributyl-

D

-lyxo-hexopyranosyl)-(1Ǟ4)-

D

-glucopyranosyl)-

D

A catalytic amount of α,αЈ-azoisobutyronitrile was added, and the

mixture was stirred at 100 °C for 2 h, then concentrated. A solution

of the residue in acetonitrile was extracted with hexane (3 ϫ), and

concentrated. Column chromatography (toluene/EtOAc, 3:1) of the

°C. A catalytic amount of α,αЈ-azoisobutyronitrile was added and

the mixture was stirred at 100 °C for 2 h, then concentrated. A

solution of the residue in acetonitrile was extracted with hexane (3

ϫ), and concentrated. Column chromatography (toluene/EtOAc,

residue yielded 41, isolated as a syrup (87 mg, 70%). TLC (toluene/ 4:1) of the residue yielded 45, isolated as a syrup (71 mg, 62%).

EtOAc, 3:1): R_fϭ 0.53. [α]²_D⁰ϭ Ϫ5 (c ϭ 1, CHCl₃). ¹H NMR TLC (toluene/EtOAc, 4:1): R_fϭ 0.45. [α]²_D⁰ϭ Ϫ1 (c ϭ 1, CHCl₃).

(300 MHz, CDCl₃): δ ϭ 0.88 (t, 3 H, octyl CH₃), 1.26 (m, 10 H, 5 ¹H NMR (300 MHz, CDCl₃): δ ϭ 0.88 (t, 3 H, octyl CH₃), 0.98

octyl CH₂), 1.50 (m, 2 H, octyl CH₂), 1.67 (s, 3 H, NHCOCH₃), (t, 3 H, NHCOCH₂CH₃), 1.25 (m, 10 H, 5 octyl CH₂), 1.50 (m, 2

1.90Ϫ2.11 (m, 2 H, 2_axЈЈ-H, 2_eqЈЈ-H), 3.12 (m, 1 H, octyl OCHH), H, octyl CH₂), 1.82Ϫ2.12 (m, 4 H, NHCOCH₂CH₃, 2_axЈЈ-H, 2_eqЈЈ-

4.13 (s, J_1,2Ͻ 1 Hz, 1 H, 1-H), 5.11 (d, J_1Ј,2Јϭ 7.7 Hz, 1 H, 1Ј-H), H), 3.10 (m, 1 H, octyl OCHH), 4.12 (s, J_1,2Ͻ 1 Hz, 1 H, 1-H),

5.61 (d, J_2Ј,NHϭ 6.8 Hz, 1 H, NH), 7.10Ϫ7.45 (m, 40 H, 8 5.15 (d, J_1Ј,2Јϭ 7.6 Hz, 1 H, 1Ј-H), 5.63 (d, J_2Ј,NHϭ 6.8 Hz, 1 H,

OCH₂C₆H₅) ppm. MS (FAB^ϩ) of C₈₄H₉₉NO₁₅(1361): found NH), 7.10Ϫ7.43 (m, 40 H, 8 OCH₂C₆H₅) ppm. MS (FAB^ϩ) of

m/z ϭ 1362 [M ϩ H]^ϩ, 1384 [M ϩ Na]^ϩ.

C₈₅H₁₀₁NO₁₅(1375): found m/z ϭ 1376 [M ϩ H]^ϩ, 1398 [M ϩ

Na]^ϩ.

Octyl

2-Deoxy-β-D-lyxo-hexopyranosyl-(1Ǟ4)-2-acetamido-2-de-

oxy-β-

D

-glucopyranosyl-(1Ǟ2)-α-

D

-mannopyranoside (7): To a solu- Octyl 2-Deoxy-β-

amido-β- -glucopyranosyl-(1Ǟ2)-α-

added 10% Pd/C (45 mg) and 6 drops of HOAc, and the mixture solution of 45 (70 mg, 51 µmol) in EtOH/EtOAc (9 mL, 1:1) were

D

-lyxo-hexopyranosyl-(1Ǟ4)-2-deoxy-2-propion-

tion of 41 (107 mg, 79 µmol) in EtOH/EtOAc (14 mL, 1:1) were

D

D-mannopyranoside (8): To a

was stirred under H₂for 21 h, then filtered through Celite, and

concentrated. Gel-filtration of the residue on a Bio-Gel P-2 col-

umn, eluted with water, and subsequent lyophilization yielded 7,

isolated as a white powder (38 mg, 75%). TLC (1-BuOH/EtOH/

HOAc/H₂O, 4:2:2:1): R_fϭ 0.29. [α]²_D⁰ϭ Ϫ9 (c ϭ 1, H₂O). High-

added 10% Pd/C (30 mg) and 6 drops of HOAc, and the mixture

was stirred under H₂overnight, then filtered through Celite, and

concentrated. Gel-filtration of the residue on a Bio-Gel P-2 col-

umn, eluted with water, and subsequent lyophilization yielded 8,

isolated as a white powder (19 mg, 57%). TLC (1-BuOH/EtOH/

resolution MS of C₂₈H₅₁NO₁₅(643.3259): calcd. 664.3156, found HOAc/H₂O, 4:2:2:1): R_fϭ 0.44. [α]²_D⁰ϭ Ϫ14 (c ϭ 1, H₂O). High-

1

664.3175 [M ϩ H]. For H and ¹³C NMR spectroscopic data, see

resolution MS of C₂₉H₅₃NO₁₅(655.3415): calcd. 678.3313, found

678.3326 [M ϩ Na]. For ¹H and ¹³C NMR spectroscopic data, see

Tables 1 and 2, respectively.

Octyl (3,4,6-Tri-O-benzyl-2-O-phenoxythiocarbonyl-β-

anosyl)-(1Ǟ4)-(3,6-di-O-benzyl-2-deoxy-2-propionamido-β-

pyranosyl)-(1Ǟ2)-3,4,6-tri-O-benzyl-α- -mannopyranoside (44): A lactopyranosyl)-(1Ǟ4)-(3,6-di-O-benzyl-2-deoxy-2-phthalimido-β-D-

D-galactopyr-

D-gluco- Octyl (4-O-Acetyl-3,6-di-O-benzyl-2-deoxy-2-phthalimido-β-D-ga-

D

solution of 37 (310 mg, 205 µmol) in 1-BuOH (28 mL), containing

glucopyranosyl)-(1Ǟ2)-3,4,6-tri-O-benzyl-α-

(47): A solution of ethyl 4-O-acetyl-3,6-di-O-benzyl-2-deoxy-2-

(46; 228 mg,

D-mannopyranoside

˚

molecular sieves 3 A (1 g), was stirred under Ar for 1 h, then 1,2-

diaminoethane (2.6 mL, 39 mmol) was added. The mixture was phthalimido-1-thio-β--galactopyranoside^[19]

stirred at 90 °C overnight, filtered through Celite, and co-concen-

trated with toluene. A solution of the residue in pyridine/propionic

anhydride (60 mL, 1:1) was stirred overnight, then co-concentrated

with toluene and EtOH. Column chromatography (toluene/EtOAc,

2:1) of the residue yielded 42 (280 mg, 94%), which was immedi-

0.40 mmol) and 13 (169 mg, 0.16 mmol) in dry CH₂Cl₂(5 mL),

containing powdered molecular sieves 4 A (0.46 g), was stirred un-

der Ar for 4 h. Then a solution of AgOTf (83 mg, 0.32 mmol) in

dry toluene (1 mL) was added and the stirring was continued for

20 min. After the addition of a solution of Br₂(0.21 mmol) in dry

˚

ately employed in the next reaction. To a solution of 42 (280 mg, CH₂Cl₂(0.5 mL) and stirring for 1 h, a second solution of AgOTf

193 µmol) in MeOH/CH₂Cl₂(75 mL, 2:1) was added NaOMe

(pH ϭ 10), and the solution was stirred at 40 °C overnight, then

(43 mg, 0.17 mmol) in dry toluene (0.5 mL) was added. When TLC

(toluene/EtOAc, 5:1) showed the reaction to be complete, the mix-

3582

 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.eurjoc.org

Eur. J. Org. Chem. 2003, 3569Ϫ3586

β-D-Galp-(1Ǟ4)-β-D-GlcpNAc-(1Ǟ2)-α-D-Manp-(1ǞO)(CH₂)₇CH₃Mimics

ture was neutralised with NEt₃, filtered through Celite, diluted with column, eluted with water, and subsequent lyophilization yielded

FULL PAPER

CH₂Cl₂, washed with aq. 10% NaHSO₃, aq. saturated NaHCO₃, 9, isolated as a white foam (24.9 mg, 56%). TLC (1-BuOH/EtOH/

and water, dried (MgSO₄), and concentrated. Column chromatog-

HOAc/H₂O, 4:2:2:1): R_fϭ 0.39. [α]²_D⁰ϭ Ϫ6 (c ϭ 0.5, H₂O). High-

raphy (toluene/EtOAc, 5:1) of the residue yielded 47, isolated as a

resolution MS of C₃₀H₅₄N₂O₁₆(698.3473): calcd. 721.3371, found

white foam (135 mg, 51%). TLC (toluene/EtOAc, 5:1): R_fϭ 0.58. 721.3351 [M ϩ Na]. For ¹H and ¹³C NMR spectroscopic data, see

[α]²_D⁰ϭ Ϫ3 (c ϭ 1, CHCl₃). ¹H NMR (300 MHz, CDCl₃): δ ϭ 0.88

(t, 3 H, octyl CH₃), 1.27 (m, 10 H, 5 octyl CH₂), 1.60 (m, 2 H,

octyl CH₂), 2.05 (s, 3 H, COCH₃), 2.98 and 3.10 (2 m, each 1 H,

Tables 1 and 2, respectively.

Octyl (3,6-Di-O-acetyl-2-deoxy-2-propionamido-4-O-propionyl-β-D-

galactopyranosyl)-(1Ǟ4)-(3,6-di-O-acetyl-2-deoxy-2-propionamido-

β- -glucopyranosyl)-(1Ǟ2)-3,4,6-tri-O-acetyl-α- -mannopyranoside

octyl OCH₂), 3.97 (s, J_1,2Ͻ 1 Hz, 1 H, 1-H), 5.13 (d, J_1Ј,2Ј

ϭ

D

7.5 Hz, 1 H, 1Ј-H), 5.37 (d, J_1ЈЈ,2ЈЈϭ 8.2 Hz, 1 H, 1ЈЈ-H), 5.64 (d,

J_3ЈЈ,4ЈЈϭ 2.9, J_4ЈЈ,5ЈЈϽ 1 Hz, 1 H, 4ЈЈ-H), 6.85Ϫ7.91 (m, 43 H, 2

Phth, 7 OCH₂C₆H₅) ppm. MS (FAB^ϩ) of C₉₃H₉₈N₂O₁₉(1546):

found m/z ϭ 1547 [M ϩ H]^ϩ, 1569 [M ϩ Na]^ϩ.

(51): A solution of 47 (138 mg, 89 µmol) in 1-BuOH (13 mL), con-

˚

taining molecular sieves 3 A (0.4 g), was stirred under Ar for

30 min, then 1,2-diaminoethane (1.3 mL, 18.6 mmol) was added.

The mixture was stirred at 90 °C overnight, filtered through Celite,

and co-concentrated with toluene and EtOH. A solution of the

residue in pyridine/propionic anhydride (10 mL, 1:1) was stirred

overnight, then co-concentrated with toluene. Column chromatog-

Octyl (2-Acetamido-4-O-acetyl-3,6-di-O-benzyl-2-deoxy-β-

topyranosyl)-(1Ǟ4)-(2-acetamido-3,6-di-O-benzyl-2-deoxy-β-

glucopyranosyl)-(1Ǟ2)-3,4,6-tri-O-benzyl-α- -mannopyranoside

D-galac-

D

-

D

(48): A solution of 47 (186 mg, 120 µmol) in 1-BuOH (17 mL), raphy (toluene/EtOAc, 5:1) of the residue yielded 50, isolated as a

˚

containing molecular sieves 3 A (0.5 g), was stirred under Ar for

30 min, then 1,2-diaminoethane (1.7 mL, 25.3 mmol) was added.

The mixture was stirred overnight at 90 °C, filtered through Celite,

and co-concentrated with toluene, EtOH and CH₂Cl₂. A solution

of the residue in pyridine/acetic anhydride (10 mL, 1:1) was stirred

overnight, then co-concentrated with toluene. Column chromatog-

raphy (toluene/EtOAc, 2:3) of the residue yielded 48, isolated as a

white foam (100 mg, 80%). TLC (CH₂Cl₂/MeOH, 9:1): R_fϭ 0.90.

To a solution of 50 (100 mg, 71 µmol) in EtOH/EtOAc (7 mL, 1:1)

were added 10% Pd/C (102 mg) and 5 drops of HOAc, and the

mixture was stirred under H₂overnight, then filtered through Celite

and concentrated. A solution of the residue in pyridine/acetic anhy-

dride (10 mL, 1:1) was stirred overnight, then co-concentrated with

toluene. Column chromatography (CH₂Cl₂/MeOH, 9:1) of the resi-

white foam (102 mg, 61%). TLC (toluene/EtOAc, 2:3): R_fϭ 0.59. due yielded 51, isolated as a syrup (71 mg, 92%). TLC (CH₂Cl₂/

[α]²_D⁰ϭ Ϫ2 (c ϭ 1, CHCl₃). ¹H NMR (300 MHz, CDCl₃): δ ϭ 0.91 MeOH, 9:1): R_fϭ 0.70. [α]²_D⁰ϭ Ϫ7 (c ϭ 1, CHCl₃). ¹H NMR

(t, 3 H, octyl CH₃), 1.29 (m, 10 H, 5 octyl CH₂), 1.53 (m, 2 H, (300 MHz, CDCl₃): δ ϭ 0.85 (t, 3 H, octyl CH₃), 1.02Ϫ1.16 (m, 9

octyl CH₂), 1.77 and 1.82 (2 s, each 3 H, 2 NHCOCH₃), 2.02 (s, 3 H, COCH₂CH₃, 2 NHCOCH₂CH₃), 1.26 (m, 10 H, 5 octyl CH₂),

H, COCH₃), 4.13 (s, J_1,2Ͻ 1 Hz, 1 H, 1-H), 5.06 (d, J_1Ј,2Јϭ 7.5 Hz, 1.55 (m, 2 H, octyl CH₂), 1.92Ϫ2.12 (m, 27 H, 7 COCH₃,

1 H, 1Ј-H), 5.12 (d, J_1ЈЈ,2ЈЈϭ 8.0 Hz, 1 H, 1ЈЈ-H), 5.59 (dd, J_3ЈЈ,4ЈЈCOCH₂CH₃, 2 NHCOCH₂CH₃), 3.39 (m, 1 H, octyl OCHH), 5.07

ϭ

2.9, J_4ЈЈ,5ЈЈϽ 1 Hz, 1 H, 4ЈЈ-H), 5.87 (d, 2 H, 2 NH), 7.18Ϫ7.39 (dd, J_2ЈЈ,3ЈЈϭ 9.9, J_3ЈЈ,4ЈЈϭ 3.3 Hz, 1 H, 3ЈЈ-H), 5.93Ϫ6.05 (m, 2 H,

(m, 35 H, 7 OCH₂C₆H₅) ppm. MS (FAB^ϩ) of C₈₁H₉₈N₂O₁₇(1370): 2 NH) ppm. MS (FAB^ϩ) of C₄₉H₇₆N₂O₂₄(1076): found m/z ϭ

found m/z ϭ 1371 [M ϩ H]^ϩ, 1393 [M ϩ Na]^ϩ.

Octyl (2-Acetamido-3,4,6-tri-O-acetyl-2-deoxy-β-

1077 [M ϩ H]^ϩ, 1099 [M ϩ Na]^ϩ.

Octyl 2-Deoxy-2-propionamido-β-

D

-galactopyrano-

-glucopyr-

-mannopyranoside (49): To a

D

-galactopyranosyl-(1Ǟ4)-2-de-

syl)-(1Ǟ4)-(2-acetamido-3,6-di-O-acetyl-2-deoxy-β-

anosyl)-(1Ǟ2)-3,4,6-tri-O-acetyl-α-

D

oxy-2-propionamido-β-D-glucopyranosyl-(1Ǟ2)-α-D-manno-

D

pyranoside (10): To a solution of 51 (71 mg, 66 µmol) in MeOH

(5 mL) was added NaOMe (pH ϭ 10), and the mixture was stirred

overnight. After neutralisation with Dowex 50 ϫ 8 (H^ϩ) and fil-

solution of 48 (102 mg, 74 µmol) in EtOH/EtOAc (7 mL, 1:1) were

added 10% Pd/C (62 mg) and 3 drops of HOAc. The mixture was

stirred under H₂overnight, then filtered through Celite, and con- tration, the solution was concentrated. Gel-filtration of the residue

centrated. A solution of the residue in pyridine/acetic anhydride

(10 mL, 1:1) was stirred overnight, then co-concentrated with tolu-

on a Bio-Gel P-2 column, eluted with water, and subsequent ly-

ophilization yielded 10, isolated as a white foam (44 mg, 91%).

ene. Column chromatography (CH₂Cl₂/MeOH, 9:1) of the residue TLC (1-BuOH/EtOH/HOAc/H₂O, 4:2:2:1): R_fϭ 0.32. [α]²_D⁰ϭ Ϫ12

yielded 49, isolated as a syrup (67 mg, 86%). TLC (CH₂Cl₂/MeOH, (c ϭ 1, H₂O). High-resolution MS of C₃₂H₅₈N₂O₁₆(726.3786):

1

9:1): R_fϭ 0.59. [α]²_D⁰ϭ Ϫ2 (c ϭ 1, CHCl₃). H NMR (300 MHz,

calcd. 749.3684, found 749.3681 [M ϩ Na]. For H and ¹³C NMR

CDCl₃): δ ϭ 0.85 (t, 3 H, octyl CH₃), 1.25 (m, 10 H, 5 octyl CH₂), spectroscopic data, see Tables 1 and 2, respectively.

1.54 (m, 2 H, octyl CH₂), 1.90, 1.91, 1.94, 1.96, 1.99, 2.02, 2.05,

2.06, 2.07, and 2.11 (10 s, each 3 H, 8 COCH₃, 2 NHCOCH₃), 3.37

1,3,4,6-Tetra-O-acetyl-2-deoxy-2-(dimethylmaleimido)-β-D-galac-

topyranose (53): To a solution of -galactosamine·HCl (52; 2.0 g,

9.3 mmol) in MeOH (69 mL) was added NaOMe (0.5 g, 9.3 mmol)

and dimethylmaleic anhydride (0.6 g, 4.8 mmol), and the mixture

was stirred at 60 °C for 30 min. Then, NEt₃(0.93 mL) and dimeth-

ylmaleic anhydride (0.6 g, 4.8 mmol) were added, and the stirring

was continued at 60 °C for 1.5 h. After concentration of the mix-

(m, 1 H, octyl OCHH), 5.05 (dd, J_2ЈЈ,3ЈЈϭ 9.9, J_3ЈЈ,4ЈЈϭ 3.3 Hz, 1

H, 3ЈЈ-H), 6.10 and 6.27 (2 d, each 1 H, 2 NH) ppm. ¹³C NMR

(75.5 MHz, CDCl₃): δ ϭ 13.9 (octyl CH₃), 20.5Ϫ20.8 (COCH₃),

23.0 and 23.1 (NHCOCH₃), 22.5, 26.0, 29.0, 29.2, 29.3, and 31.6

(6 octyl CH₂), 51.6 and 54.1 (C-2Ј, C-2ЈЈ), 61.0, 62.6, 62.7, and 68.2

(C-6, C-6Ј, C-6ЈЈ, octyl OCH₂), 97.4, 99.0, and 100.5 (C-1, C-1Ј,

C-1ЈЈ), 169.3, 170.0, 170.1, 170.2 (3 C), 170.3, 170.5 (2 C), and

170.7 (COCH₃, NHCOCH₃) ppm. MS (FAB^ϩ) of C₄₆H₇₀N₂O₂₄

(1034): found m/z ϭ 1035 [M ϩ H]^ϩ, 1057 [M ϩ Na]^ϩ.

ture,

a solution of the residue in pyridine/acetic anhydride

(28.5 mL, 2:1) was stirred overnight, then co-concentrated with

toluene. A solution of the residue in CH₂Cl₂(50 mL) was washed

with aq. 3% HCl, aq. saturated NaHCO₃, and water, dried

Octyl

2-Acetamido-2-deoxy-β-D-galactopyranosyl-(1Ǟ4)-2-acet- (MgSO₄), filtered, and concentrated. Column chromatography

amido-2-deoxy-β-

D

-glucopyranosyl-(1Ǟ2)-α-

D

-mannopyranoside (hexane/EtOAc, 1:1) of the residue yielded 53, isolated as a white

(9): To a solution of 49 (65 mg, 63 µmol) in MeOH (4 mL) was

added NaOMe (pH ϭ 10), and the mixture was stirred overnight.

After neutralisation with Dowex 50 ϫ 8 (H^ϩ) and filtration, the

solution was concentrated. Gel-filtration through a Bio-Gel P-2

foam (1.3 g, 31%). TLC (hexane/EtOAc, 1:1): R_fϭ 0.30. [α]²_D⁰

ϭ

ϩ48 (c ϭ 1, CHCl₃). ¹H NMR (300 MHz, CDCl₃): δ ϭ 1.91, 1.98,

2.04, and 2.17 (4 s, 3, 6, 6, 3 H, 2 CH₃, 4 COCH₃), 4.41 (dd, J_1,2ϭ

8.9, J_2,3ϭ 11.4 Hz, 1 H, 2-H), 5.47 (d, J_3,4ϭ 3.4, J_4,5Ͻ 1 Hz, 1

Eur. J. Org. Chem. 2003, 3569Ϫ3586

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 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

3583

J. P. Kamerling et al.

FULL PAPER

H, 4-H), 5.77 (dd, 1 H, 3-H), 6.28 (d, 1 H, 1-H) ppm. MS Octyl (2-Deoxy-2-propionamido-3,4,6-tri-O-propionyl-β-

D

-galacto-

pyranosyl)-(1Ǟ4)-(3,6-di-O-benzyl-2-deoxy-2-phthalimido-β-

glucopyranosyl)-(1Ǟ2)-3,4,6-tri-O-benzyl-α- -mannopyranoside

(MALDI-TOF) of C₂₀H₂₅NO₁₁(455): found m/z ϭ 478 [M ϩ

D-

Na]^ϩ, 494 [M ϩ K]^ϩ.

D

(57): To a solution of 56 (364 mg, 255 µmol) in dioxane/water

(14.4 mL, 4:1) was added NaOH (287 mg, 7.18 mmol), and the

mixture was stirred at room temperature overnight. Then, the pH

was adjusted to 3 using aq. 4  HCl, and the stirring was continued

overnight. The mixture was neutralised with solid K₂CO₃, and co-

concentrated with toluene. A solution of the residue in pyridine/

propionic anhydride (20 mL, 1:1) was stirred overnight, then co-

concentrated with toluene. The residue was dissolved in CH₂Cl₂,

and the solution was washed with aq. 1  HCl, aq. saturated

NaHCO₃, and water, dried (MgSO₄), filtered, and concentrated.

Column chromatography (hexane/EtOAc, 1:2) of the residue

yielded 57, isolated as a colourless syrup (116 mg, 33%). TLC (hex-

3,4,6-Tri-O-acetyl-2-deoxy-2-(dimethylmaleimido)-D-galacto-

pyranose (54): To a solution of 53 (0.78 g, 1.7 mmol) in DMF

(6 mL) was added hydrazinium acetate (0.19 g, 2.1 mmol). The

mixture was stirred at room temperature for 1 h, diluted with

EtOAc, washed with cold aq. saturated NaHCO₃and water, dried

(MgSO₄), filtered, and concentrated. Column chromatography

(toluene/EtOAc, 1:1) of the residue yielded 54, isolated as a white

foam (0.43 g, 61%). TLC (toluene/EtOAc, 1:1): R_fϭ 0.37. ¹H

NMR β-anomer (300 MHz, CDCl₃): δ ϭ 1.89, 1.96, 2.05, and 2.17

(4 s, 3, 6, 3, 3 H, 2 CH₃, 3 COCH₃), 4.83 (dd, J_1,2ϭ 7.0, J_2,3

11.5 Hz, 1 H, 2-H), 5.33 (d, 1 H, 1-H), 5.43 (d, J_3,4ϭ 3.3, J_4,5

ϭ

1

ane/EtOAc, 1:1): R_fϭ 0.45. [α]²_D⁰ϭ ϩ9 (c ϭ 1, CHCl₃). H NMR

Ͻ

1 Hz, 1 H, 4-H), 5.65 (dd, 1 H, 3-H) ppm. MS (MALDI-TOF) of

(300 MHz, CDCl₃): δ ϭ 0.87 (t, 3 H, octyl CH₃), 1.02Ϫ1.14 (m,

12 H, 3 COCH₂CH₃, NHCOCH₂CH₃), 1.24 (m, 10 H, 5 octyl

CH₂), 1.44 (m, 2 H, octyl CH₂), 1.90 (m, 2 H, NHCOCH₂CH₃),

2.20Ϫ2.36 (m, 6 H, 3 COCH₂CH₃), 3.20 (m, 1 H, octyl OCHH),

5.29 (d, J_4ЈЈ,5ЈЈϽ 1 Hz, 1 H, 4ЈЈ-H), 6.99Ϫ7.41 (m, 29 H, Phth, 5

OCH₂C₆H₅) ppm. ¹³C NMR (75.5 MHz, CDCl₃): δ ϭ 8.7, 8.8, 9.0,

and 9.6 (3 COCH₂CH₃, NHCOCH₂CH₃), 13.9 (octyl CH₃), 22.5,

25.9, 27.2 (2 C), 29.0, 29.2 (2 C), 29.5 (2 C), and 31.6 (6 octyl CH₂,

3 COCH₂CH₃, NHCOCH₂CH₃), 50.8 (2 C) (C-2Ј, C-2ЈЈ), 60.8,

67.4, 69.0, 69.6, 70.5, 72.7 (2 C), 73.8, and 74.0 (C-6, C-6Ј, C-6ЈЈ,

5 OCH₂C₆H₅, octyl OCH₂), 63.9, 66.1, 70.5 (2 C), 71.4, 74.7, 74.8,

77.0, 78.1, and 82.1 (C-2, C-3, C-4, C-5, C-3Ј, C-4Ј, C-5Ј, C-3ЈЈ, C-

4ЈЈ, C-5ЈЈ), 97.5, 100.8, and 101.7 (C-1, C-1Ј, C-1ЈЈ), 173.4 and

173.5 (COCH₂CH₃, NHCOCH₂CH₃) ppm. MS (MALDI-TOF) of

C₈₁H₉₈N₂O₂₀(1419): found m/z ϭ 1442 [M ϩ Na]^ϩ, 1458 [M ϩ

K]^ϩ.

C₁₈H₂₃NO₁₀(413): found m/z ϭ 436 [M ϩ Na]^ϩ, 452 [M ϩ K]^ϩ.

3,4,6-Tri-O-acetyl-2-deoxy-2-(dimethylmaleimido)-β-D-galacto-

pyranosyl trichloroacetimidate (55): To a solution of 54 (0.30 g,

0.73 mmol) in CH₂Cl₂(6 mL) were added trichloroacetonitrile

(1.05 g, 7.3 mmol) and 1,8-diazabicyclo[5.4.0]undec-7-ene (29 mg,

0.19 mmol), and the mixture was stirred for 30 min at room tem-

perature, then concentrated. Column chromatography (toluene/

EtOAc, 1:1) of the residue yielded 55, isolated as a white foam

(0.24 g, 59%). TLC (toluene/EtOAc, 1:1): R_fϭ 0.56. [α]²_D⁰ϭ ϩ36

1

(c ϭ 1, CHCl₃). H NMR (300 MHz, CDCl₃): δ ϭ 1.93, 2.05, and

2.20 (3 s, 9, 3, 3 H, 2 CH₃, 3 COCH₃), 4.58 (dd, J_1,2ϭ 8.6, J_2,3

ϭ

11.5 Hz, 1 H, 2-H), 5.50 (d, J_3,4ϭ 3.4, J_4,5Ͻ 1 Hz, 1 H, 4-H), 5.78

(dd, 1 H, 3-H), 6.40 (d, 1 H, 1-H), 8.67 (s, 1 H, NH) ppm. ¹³C

NMR (75.5 MHz, CDCl₃): δ ϭ 8.6 (CH₃), 20.3Ϫ20.5 (COCH₃),

49.9 (C-2), 60.8 (C-6), 66.2, 67.6, and 71.6 (C-3, C-4, C-5), 93.9

(C-1), 137.3 (CϭC NDMM), 160.4 (CONHCCl₃), 169.5Ϫ170.2

(COCH₃, CO NDMM).

Octyl (3,4,6-Tri-O-acetyl-2-deoxy-2-propionamido-β-

anosyl)-(1Ǟ4)-(2-acetamido-3,6-di-O-benzyl-2-deoxy-β-

anosyl)-(1Ǟ2)-3,4,6-tri-O-benzyl-α- -mannopyranoside (58):

D-galactopyr-

D

-glucopyr-

D

A

solution of 57 (116 mg, 84.2 µmol) in 1-BuOH (22 mL) was stirred

at room temperature for 30 min, then 1,2-diaminoethane (1.2 mL,

18 mmol) was added. The solution was stirred at 90 °C overnight,

then co-concentrated with toluene. A solution of the residue in

pyridine/acetic anhydride (9.6 mL, 1:1) was stirred overnight, then

co-concentrated with toluene. Column chromatography (toluene/

EtOAc, 2:3) of the residue yielded 58, isolated as a colourless syrup

(78 mg, 74%). TLC (toluene/EtOAc, 2:3): R_fϭ 0.41. [α]²_D⁰ϭ ϩ3

(c ϭ 1, CHCl₃). ¹H NMR (300 MHz, CDCl₃): δ ϭ 0.88 (t, 3 H,

octyl CH₃), 1.06 (t, 3 H, NHCOCH₂CH₃), 1.26 (m, 10 H, 5 octyl

CH₂), 1.57 (m, 2 H, octyl CH₂), 1.78 (s, 3 H, NHCOCH₃), 1.96,

2.01, and 2.07 (3 s, each 3 H, 3 COCH₃), 2.05 (m, 2 H,

NHCOCH₂CH₃), 3.34 (m, 1 H, octyl OCHH), 4.09 (s, J_1,2Ͻ 1 Hz,

1 H, 1-H), 4.91 (dd, J_2ЈЈ,3ЈЈϭ 11.3, J_3ЈЈ,4ЈЈϭ 3.2 Hz, 1 H, 3ЈЈ-H),

5.25 (d, J_4ЈЈ,5ЈЈϽ 1 Hz, 1 H, 4ЈЈ-H), 5.91 (d, 1 H, NH), 7.20Ϫ7.39

(m, 25 H, 5 OCH₂C₆H₅) ppm. ¹³C NMR (75.5 MHz, CDCl₃): δ ϭ

9.6 (NHCOCH₂CH₃), 13.9 (octyl CH₃), 20.4Ϫ20.5 (COCH₃), 23.1

(NHCOCH₃), 22.5, 26.0, 29.0Ϫ29.8, and 31.6 (octyl CH₂,

NHCOCH₂CH₃), 50.9 and 55.7 (C-2Ј, C-2ЈЈ), 61.0, 67.7, 69.2, 71.1,

73.1 (2 C), 73.3, 73.5, and 74.9 (C-6, C-6Ј, C-6ЈЈ, 5 OCH₂C₆H₅,

octyl OCH₂), 66.3, 70.1, 70.4, 71.5, 73.6, 74.1, 74.5, 76.2, 77.0, and

78.4 (C-2, C-3, C-4, C-5, C-3Ј, C-4Ј, C-5Ј, C-3ЈЈ, C-4ЈЈ, C-5ЈЈ), 97.5,

97.9, and 100.2 (C-1, C-1Ј, C-1ЈЈ), 170.1Ϫ171.0 (COCH₃,

NHCOCH₃), 173.8 (NHCOCH₂CH₃) ppm.

Octyl (3,4,6-Tri-O-acetyl-2-deoxy-2-dimethylmaleimido-β-

pyranosyl)-(1Ǟ4)-(3,6-di-O-benzyl-2-deoxy-2-phthalimido-β-

pyranosyl)-(1Ǟ2)-3,4,6-tri-O-benzyl-α- -mannopyranoside (56): A

D

-galacto-

D-gluco-

D

solution of 55 (147 mg, 264 µmol) and 13 (182 mg, 176 µmol) in

˚

dry CH₂Cl₂(2.2 mL), containing molecular sieves 4 A (30 mg), was

stirred under Ar at room temperature for 45 min. After cooling to

0 °C, a solution of TMSOTf (2.64 µmol) in dry CH₂Cl₂(26.4 µL)

was added. When TLC (hexane/EtOAc, 1:1) showed the reaction

to be complete, the mixture was neutralised with NEt₃and concen-

trated. Column chromatography (hexane/EtOAc, 1:1) of the residue

yielded 56, isolated as a colourless syrup (212 mg, 84%). TLC (hex-

ane/EtOAc, 1:1): R_fϭ 0.59. [α]²_D⁰ϭ ϩ6 (c ϭ 1, CHCl₃). H NMR

1

(300 MHz, CDCl₃): δ ϭ 0.86 (t, 3 H, octyl CH₃), 1.23 (m, 10 H, 5

octyl CH₂), 1.36 (m, 2 H, octyl CH₂), 1.88, 1.93, 2.02, and 2.04 (4

s, 3, 3, 6, 3 H, 2 CH₃, 3 COCH₃), 2.91 and 3.13 (2 m, each 1 H,

octyl OCH₂), 5.32 (d, J_3ЈЈ,4ЈЈϭ 3.4, J_4ЈЈ,5ЈЈϽ 1 Hz, 1 H, 4ЈЈ-H), 5.32

(d, J_1Ј,2Јϭ 8.2 Hz, 1 H, 1Ј-H), 5.61 (dd, J_2ЈЈ,3ЈЈϭ 11.5 Hz, 1 H, 3ЈЈ-

H), 6.90Ϫ7.34 (m, 25 H, 5 OCH₂C₆H₅), 7.55 and 7.68 (2 m, each

2 H, Phth) ppm. ¹³C NMR (75.5 MHz, CDCl₃): δ ϭ 8.9 (CH₃),

14.0 (octyl CH₃), 20.4Ϫ20.5 (COCH₃), 22.5, 25.9, 29.1, 29.2, 29.6,

and 31.7 (octyl CH₂), 51.8 and 55.3 (C-2Ј, C-2ЈЈ), 60.9, 67.6, 68.7,

69.8, 70.5, 72.7, 73.1, 73.7, and 74.7 (C-6, C-6Ј, C-6ЈЈ,

5

OCH₂C₆H₅, octyl OCH₂), 66.6, 67.8, 70.5, 72.5, 73.3, 74.6, 74.7,

75.9, 76.3, and 77.6 (C-2, C-3, C-4, C-5, C-3Ј, C-4Ј, C-5Ј, C-3ЈЈ, C-

4ЈЈ, C-5ЈЈ), 96.6 (2 C) and 97.1 (C-1, C-1Ј, C-1ЈЈ), 169.6Ϫ170.2 Octyl (3,4,6-Tri-O-acetyl-2-deoxy-2-propionamido-β-

D

-galactopyr-

anosyl)-(1Ǟ4)-(2-acetamido-3,6-di-O-acetyl-2-deoxy-β- -glucopyr-

-mannopyranoside (59): To a

(COCH₃, CO NDMM) ppm. MS (MALDI-TOF) of C₈₁H₉₂N₂O₂₁

D

(1429): found m/z ϭ 1452 [M ϩ Na]^ϩ, 1468 [M ϩ K]^ϩ.

anosyl)-(1Ǟ2)-3,4,6-tri-O-acetyl-α-

D

3584

 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.eurjoc.org

Eur. J. Org. Chem. 2003, 3569Ϫ3586

β-D-Galp-(1Ǟ4)-β-D-GlcpNAc-(1Ǟ2)-α-D-Manp-(1ǞO)(CH₂)₇CH₃Mimics

FULL PAPER

solution of 58 (78 mg, 63 µmol) in MeOH/CH₂Cl₂(4.0 mL, 1:1)

was added NaOMe (pH ϭ 9), and the mixture was stirred over-

night. After neutralisation with Dowex 50 ϫ 8 (H^ϩ) and filtration,

co-concentrated with toluene. A solution of the residue in pyridine/

acetic anhydride (5.3 mL, 2:1) was stirred overnight, then co-con-

centrated with toluene. The residue was dissolved in CH₂Cl₂, and

the solution was concentrated. To a solution of the residue in the solution was washed with aq. 1  HCl, aq. saturated NaHCO₃,

EtOH/EtOAc (6.0 mL, 1:1) were added 10% Pd/C (52 mg) and 3 and water, dried (MgSO₄), filtered, and concentrated. Column

drops of HOAc, and the mixture was stirred under H₂overnight, chromatography (hexane/EtOAc, 1:2) of the residue yielded 60, iso-

then filtered, and concentrated. A solution of the residue in pyri-

dine/acetic anhydride (9.0 mL, 1:1) was stirred overnight, then co-

lated as a colourless syrup (23 mg, 48%). TLC (hexane/EtOAc, 1:2):

R_fϭ 0.36. [α]²_D⁰ϭ ϩ4 (c ϭ 1, CHCl₃). ¹H NMR (300 MHz,

concentrated with toluene. Column chromatography (CH₂Cl₂/ace- CDCl₃): δ ϭ 0.87 (t, 3 H, octyl CH₃), 1.22 (m, 10 H, 5 octyl CH₂),

tone, 2:1) of the residue yielded 59, isolated as a white solid (48 mg,

1.43 (m, 2 H, octyl CH₂), 1.76 (s, 3 H, NHCOCH₃), 1.97 and 2.05

(2 s, 3, 6 H, 3 COCH₃), 2.99 and 3.17 (2 m, each 1 H, octyl OCH₂),

72%). TLC (CH₂Cl₂/acetone, 2:1): R_fϭ 0.32. [α]²_D⁰ϭ Ϫ18 (c ϭ 1,

CHCl₃). ¹H NMR (500 MHz, CDCl₃; 2D TOCSY, ROESY, 5.24 (d, J_4ЈЈ,5ЈЈϽ 1 Hz, 1 H, 4ЈЈ-H), 6.85Ϫ7.62 (m, 29 H, Phth, 5

HSQC):

δ

ϭ

0.89 (t,

3

H, octyl CH₃), 1.09 (t,

3

H, OCH₂C₆H₅) ppm. ¹³C NMR (75.5 MHz, CDCl₃): δ ϭ 14.0 (octyl

CH₃), 20.5Ϫ20.6 (COCH₃), 23.1 (NHCOCH₃), 22.5, 25.9, 29.0,

NHCOCH₂CH₃), 1.29 (m, 10 H, 5 octyl CH₂), 1.59 (m, 2 H, octyl

CH₂), 1.96, 1.97, 2.00, 2.03, 2.06, 2.09, 2.11, and 2.15 (8 s, 3, 3, 3, 29.2, 29.5, and 31.7 (octyl CH₂), 50.7 (C-2ЈЈ), 55.5 (C-2Ј), 96.9 (2

3, 3, 6, 3,

3

H,

8

COCH₃, NHCOCH₃), 2.16 (m,

2

H, C) and 100.8 (C-1, C-1Ј, C-1ЈЈ), 169.6Ϫ170.2 (COCH₃,

NHCOCH₃) ppm.

NHCOCH₂CH₃), 3.41 and 3.63 (2 ddd, each 1 H, octyl OCH₂),

3.63Ϫ3.79 (m, 3 H, 2Ј-H, 4Ј-H, 5Ј-H), 3.84Ϫ3.90 (m, 2 H, 5-H,

5ЈЈ-H), 3.92 (dd, J_1ЈЈ,2ЈЈϭ 8.3, J_2ЈЈ,3ЈЈϭ 11.2 Hz, 1 H, 2ЈЈ-H),

4.05Ϫ4.14 (m, 4 H, 2-H, 6a-H, 6ЈЈa-H, 6ЈЈb-H), 4.18 (dd, J_5,6b

5.8, J_6a,6bϭ 12.1 Hz, 1 H, 6b-H), 4.29 (dd, J_5Ј,6Јbϭ 5.0, J_6Јa,6Јb

Octyl (2-Acetamido-2-deoxy-3,4,6-tri-O-propionyl-β-

anosyl)-(1Ǟ4)-(3,6-di-O-benzyl-2-deoxy-2-propionamido-β-

glucopyranosyl)-(1Ǟ2)-3,4,6-tri-O-benzyl-α- -mannopyranoside

D-galactopyr-

D

-

ϭ

D

ϭ

(61): A solution of 60 (54 mg, 40 µmol) in 1-BuOH (5.6 mL) was

stirred for 30 min at room temperature, then 1,2-diaminoethane

(0.56 mL, 8.4 mmol) was added. The mixture was stirred overnight

at 90 °C, then co-concentrated with toluene. A solution of the resi-

due in pyridine/propionic anhydride (4.4 mL, 1:1) was stirred over-

night, then co-concentrated with toluene. Column chromatography

(toluene/EtOAc, 2:3) of the residue yielded 61, isolated as a colour-

less syrup (42 mg, 80%). TLC (toluene/EtOAc, 2:3): R_fϭ 0.39.

[α]²_D⁰ϭ ϩ1 (c ϭ 1, CHCl₃). ¹H NMR (300 MHz, CDCl₃): δ ϭ 0.88

(t, 3 H, octyl CH₃), 1.01 (m, 3 H, NHCOCH₂CH₃), 1.04Ϫ1.16 (m,

9 H, 3 COCH₂CH₃), 1.26 (m, 10 H, 5 octyl CH₂), 1.51 (m, 2 H,

11.8 Hz, 1 H, 6Јb-H), 4.34 (dd, J_5Ј,6Јaϭ 3.0 Hz, 1 H, 6Јa-H), 4.66

(d, 1 H, 1ЈЈ-H), 4.72 (d, J_1,2Ͻ 1 Hz, 2 H, 1-H, 1Ј-H), 5.10 (dd,

J_2,3ϭ 3.4, J_3,4ϭ 10.1 Hz, 1 H, 3-H), 5.20 (dd, J_3ЈЈ,4ЈЈϭ 3.4 Hz, 1

H, 3ЈЈ-H), 5.22 (t, 1 H, 4-H), 5.30 (t, 1 H, 3Ј-H), 5.32 (d, J_4ЈЈ,5ЈЈ

Ͻ

1 Hz, 1 H, 4ЈЈ-H), 5.87 (d, J_2ЈЈ,NHЈЈϭ 8.7 Hz, 1 H, NHЈЈ), 5.94 (d,

J_2Ј,NHЈϭ 8.6 Hz, 1 H, NHЈ) ppm. ¹³C NMR (75.5 MHz, CDCl₃):

δ ϭ 9.5 (NHCOCH₂CH₃), 13.9 (octyl CH₃), 20.4Ϫ20.8 (COCH₃),

23.0 (NHCOCH₃), 22.5, 26.0, 29.1Ϫ29.6, and 31.7 (octyl CH₂,

NHCOCH₂CH₃), 51.5 (C-2ЈЈ), 53.9 (C-2Ј), 61.0, 62.7 (2 C), and

68.3 (C-6, C-6Ј, C-6ЈЈ, octyl OCH₂), 66.1, 66.3, 68.4, 69.6, 70.2,

70.5, 71.2, 72.7, 74.5, and 75.2 (C-2, C-3, C-4, C-5, C-3Ј, C-4Ј, C-

5Ј, C-3ЈЈ, C-4ЈЈ, C-5ЈЈ), 97.4 and 99.1 (C-1, C-1Ј), 100.5 (C-1ЈЈ),

169.3Ϫ170.7 (COCH₃, NHCOCH₃), 174.2 (NHCOCH₂CH₃) ppm.

MS (MALDI-TOF) of C₄₇H₇₂N₂O₂₄(1048): found m/z ϭ 1049 [M

ϩ H]^ϩ, 1071 [M ϩ Na]^ϩ, 1087 [M ϩ K]^ϩ.

octyl CH₂), 1.75 (s,

3 H, NHCOCH₃), 1.96 (m, 2 H,

NHCOCH₂CH₃), 2.25 (m, 6 H, 3 COCH₂CH₃), 3.33 (m, 1 H, octyl

OCHH), 4.09 (s, J_1,2Ͻ 1 Hz, 1 H, 1-H), 5.24 (d, J_4ЈЈ,5ЈЈϽ 1 Hz, 1

H, 4ЈЈ-H), 7.18Ϫ7.41 (m, 25 H, 5 OCH₂C₆H₅) ppm. ¹³C NMR

(75.5 MHz, CDCl₃): δ ϭ 8.7, 8.8, 9.0, and 9.4 (COCH₂CH₃,

Octyl 2-Deoxy-2-propionamido-β-D-galactopyranosyl-(1Ǟ4)-2-acet- NHCOCH₂CH₃), 14.0 (octyl CH₃), 23.1 (NHCOCH₃), 22.5, 26.0,

amido-2-deoxy-β- -glucopyranosyl-(1Ǟ2)-α-

D

-mannopyranoside

27.2, 29.1Ϫ29.5, and 31.7 (octyl CH₂, COCH₂CH₃,

NHCOCH₂CH₃), 51.2 (C-2ЈЈ), 56.5 (C-2Ј), 97.6, 98.0, and 100.2

(C-1, C-1Ј, C-1ЈЈ), 169.8 (NHCOCH₃), 173.5Ϫ174.5 (COCH₂CH₃,

(11): To a solution of 59 (33 mg, 31 µmol) in MeOH/CH₂Cl₂

(2.0 mL, 1:1) was added NaOMe (pH ϭ 10), and the mixture was

stirred overnight. After neutralisation with Dowex 50 ϫ 8 (H^ϩ) NHCOCH₂CH₃) ppm.

and filtration, the solution was concentrated. The residue was ap-

plied on a C18-Bakerbond spe^tmcolumn (500 mg), eluted with

water and MeOH, and the MeOH fractions were concentrated.

Gel-filtration of the residue on a HW-40S Toyopearl column, eluted

with aq. 5 m NH₄OAc, and subsequent lyophilization yielded 11,

isolated as a white foam (17 mg, 77%). TLC (MeOH/CH₂Cl₂, 3:1):

R_fϭ 0.65. [α]²_D⁰ϭ Ϫ9 (c ϭ 1, H₂O). High-resolution MS of

C₃₁H₅₆N₂O₁₆(712.3630): calcd. 735.3528, found 735.3507 [M ϩ

Na]. For ¹H and ¹³C NMR spectroscopic data, see Tables 1 and

2, respectively.

Octyl (2-Acetamido-3,4,6-tri-O-acetyl-2-deoxy-β-

syl)-(1Ǟ4)-(3,6-di-O-acetyl-2-deoxy-2-propionamido-β-

anosyl)-(1Ǟ2)-3,4,6-tri-O-acetyl-α- -mannopyranoside (62): To a

D

-galactopyrano-

D

-glucopyr-

D

solution of 61 (42 mg, 32 µmol) in MeOH/CH₂Cl₂(2.0 mL, 1:1)

was added NaOMe (pH ϭ 9), and the mixture was stirred over-

night. After neutralisation with Dowex 50 ϫ 8 (H^ϩ) and filtration,

the solution was concentrated. To a solution of the residue in

EtOH/EtOAc (3.0 mL, 1:1) were added 10% Pd/C (27 mg) and 2

drops of HOAc, and the mixture was stirred under H₂overnight,

then filtered through Celite, and concentrated. A solution of the

residue in pyridine/acetic anhydride (5.0 mL, 1:1) was stirred over-

night, then co-concentrated with toluene. Column chromatography

(CH₂Cl₂/acetone, 1:1) of the residue yielded 62, isolated as a white

Octyl (2-Acetamido-3,4,6-tri-O-acetyl-2-deoxy-β-

syl)-(1Ǟ4)-(3,6-di-O-benzyl-2-deoxy-2-phthalimido-β-

anosyl)-(1Ǟ2)-3,4,6-tri-O-benzyl-α- -mannopyranoside (60): To a

D-galactopyrano-

D

-glucopyr-

D

solution of 56 (50 mg, 35 µmol) in dioxane/water (1.75 mL, 4:1)

was added NaOH (35 mg, 0.88 mmol), and the mixture was stirred

solid (22 mg, 67%). TLC (CH₂Cl₂/acetone, 1:1): R_fϭ 0.53. [α]²_D⁰

Ϫ21 (c ϭ 1, CHCl₃). ¹H NMR (500 MHz, CDCl₃; 2D TOCSY,

ϭ

at room temperature overnight. Then the pH was adjusted to 3 ROESY, HSQC): δ ϭ 0.89 (t, 3 H, octyl CH₃), 1.11 (t, 3 H,

using aq. 4  HCl, and the stirring was continued overnight. Sub-

sequently, the solution was made alkaline with aq. 1  NaOH fol-

NHCOCH₂CH₃), 1.28 (m, 10 H, 5 octyl CH₂), 1.59 (m, 2 H, octyl

CH₂), 1.94, 1.98, 2.01, 2.03, 2.06, 2.08, 2.12, and 2.14 (8 s, 3, 3, 3,

lowed by the addition of AcCl (43.8 µL). After stirring at pH ϭ 10 3, 3, 6, 3,

for 45 min, the pH was lowered to 3 using aq. 4  HCl, and the NHCOCH₂CH₃), 3.40 and 3.63 (2 ddd, each 1 H, octyl OCH₂),

mixture was stirred overnight, neutralised with solid K₂CO₃, and 3.65Ϫ3.73 (m, 3 H, 2Ј-H, 4Ј-H, 5Ј-H), 3.83Ϫ3.91 (m, 3 H, 5-H,

3 H, 8 COCH₃, NHCOCH₃), 2.17 (m, 2 H,

Eur. J. Org. Chem. 2003, 3569Ϫ3586

www.eurjoc.org

 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

3585

J. P. Kamerling et al.

FULL PAPER

[7]

[8]

G. Baisch, R. Öhrlein, M. Streiff, B. Ernst, Bioorg. Med. Chem.

Lett. 1996, 6, 755Ϫ758.

2ЈЈ-H, 5ЈЈ-H), 4.05 (dd, J_5,6aϭ 2.8, J_6a,6bϭ 11.9 Hz, 1 H, 6a-H),

4.07Ϫ4.13 (m, 3 H, 2-H, 6ЈЈa-H, 6ЈЈb-H), 4.17 (dd, J_5,6bϭ 5.7 Hz,

1 H, 6b-H), 4.27 (dd, J_5Ј,6Јbϭ 5.2, J_6Јa,6Јbϭ 12.0 Hz, 1 H, 6Јb-H),

4.33 (dd, J_5Ј,6Јaϭ 2.6 Hz, 1 H, 6Јa-H), 4.66 (d, J_1ЈЈ,2ЈЈϭ 8.1 Hz, 1

H, 1ЈЈ-H), 4.67 (s, J_1,2Ͻ 1 Hz, 1 H, 1-H), 4.77 (d, J_1Ј,2Јϭ 7.6 Hz,

1 H, 1Ј-H), 5.08 (dd, J_2,3ϭ 3.5, J_3,4ϭ 10.1 Hz, 1 H, 3-H), 5.19

(dd, J_2ЈЈ,3ЈЈϭ 10.8, J_3ЈЈ,4ЈЈϭ 3.3 Hz, 1 H, 3ЈЈ-H), 5.21 (t, 1 H, 4-H),

´

A. Lubineau, K. Basset-Carpentier, C. Auge, Carbohydr. Res.

1997, 300, 161Ϫ167.

[9]

C. H. Hokke, J. G. M. van der Ven, J. P. Kamerling, J. F. G.

Vliegenthart, Glycoconjugate J. 1993, 10, 82Ϫ90.

J. A. L. M. van Dorst, C. J. van Heusden, A. F. Voskamp, J. P.

Kamerling, J. F. G. Vliegenthart, Carbohydr. Res. 1996, 291,

63Ϫ83.

J. A. L. M. van Dorst, C. J. van Heusden, J. M. Tikkanen,

J. P. Kamerling, J. F. G. Vliegenthart, Carbohydr. Res. 1997,

297, 209Ϫ227.

[10]

5.32 (d, J_4ЈЈ,5ЈЈϽ 1 Hz, 1 H, 4ЈЈ-H), 5.38 (dd, J_2Ј,3Јϭ 8.2, J_3Ј,4Ј

ϭ

[11]

9.4 Hz, 1 H, 3Ј-H), 5.68 (d, J_2Ј,NHЈϭ 8.3 Hz, 1 H, NHЈ), 5.76 (d,

J_2ЈЈ,NHЈЈϭ 8.8 Hz, 1 H, NHЈЈ) ppm. ¹³C NMR (75.5 MHz, CDCl₃):

δ ϭ 9.5 (NHCOCH₂CH₃), 13.9 (octyl CH₃), 20.4Ϫ20.7 (COCH₃),

23.1 (NHCOCH₃), 22.5, 25.9, 29.1 (2 C), 29.3, 29.5, and 31.6 (octyl

CH₂, NHCOCH₂CH₃), 51.6 (C-2ЈЈ), 54.4 (C-2Ј), 61.0, 62.6, 62.7,

and 68.2 (C-6, C-6Ј, C-6ЈЈ, octyl OCH₂), 66.1, 66.3, 68.5, 69.6, 70.2,

70.5, 71.3, 72.6, 74.5, and 75.2 (C-2, C-3, C-4, C-5, C-3Ј, C-4Ј, C-

5Ј, C-3ЈЈ, C-4ЈЈ, C-5ЈЈ), 97.3 and 99.1 (C-1, C-1Ј), 100.3 (C-1ЈЈ),

169.3Ϫ170.8 (COCH₃, NHCOCH₃), 173.9 (NHCOCH₂CH₃) ppm.

[12]

[13]

J. A. L. M. van Dorst, A. F. Voskamp, J. P. Kamerling, J. F. G.

Vliegenthart, Liebigs Ann./Recueil 1997, 1227Ϫ1233.

J. A. L. M. van Dorst, J. M. Tikkanen, C. H. Krezdorn, M. B.

Streiff, E. G. Berger, J. A. van Kuik, J. P. Kamerling, J. F. G.

Vliegenthart, Eur. J. Biochem. 1996, 242, 674Ϫ681.

M. C. Galan, A. P. Venot, J. Glushka, A. Imberty, G. J. Boons,

J. Am. Chem. Soc. 2002, 124, 5964Ϫ5973.

[14]

[15]

[16]

[17]

[18]

[19]

[20]

[21]

[22]

R. R. Schmidt, M. Stumpp, Liebigs Ann. Chem. 1983,

Octyl 2-Acetamido-2-deoxy-β-

D

-galactopyranosyl-(1Ǟ4)-2-deoxy-2-

1249Ϫ1256.

propionamido-β- -glucopyranosyl)-(1Ǟ2)-α-

D

-mannopyranoside

O. Kanie, S. C. Crawley, M. M. Palcic, O. Hindsgaul, Car-

bohydr. Res. 1993, 243, 139Ϫ164.

S. Nilsson, H. Lönn, T. Norberg, Glycoconjugate J. 1989, 6,

(12): To a solution of 62 (22 mg, 21 µmol) in MeOH/CH₂Cl₂

(2.0 mL, 1:1) was added NaOMe (pH ϭ 9), and the mixture was

stirred overnight. After neutralisation with Dowex 50 ϫ 8 (H^ϩ)

and filtration, the solution was concentrated. The residue was ap-

plied on a C18-Bakerbond spe^tmcolumn (500 mg), eluted with

water and MeOH, and the MeOH fractions were concentrated.

Gel-filtration of the residue on a HW-40S Toyopearl column, eluted

with aq. 5 m NH₄OAc, and subsequent lyophilization yielded 12,

isolated as a white foam (13 mg, 84%). TLC (MeOH/CH₂Cl₂, 3:1):

R_fϭ 0.65. [α]²_D⁰ϭ Ϫ8 (c ϭ 0.5; H₂O). High-resolution MS of

C₃₁H₅₆N₂O₁₆(712.3630): calcd. 735.3528, found 735.3515 [M ϩ

Na]. For ¹H and ¹³C NMR spectroscopic data, see Tables 1 and

2, respectively.

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Received May 14, 2003

3586

 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

www.eurjoc.org

Eur. J. Org. Chem. 2003, 3569Ϫ3586

Article Doi

Synthesis of β-D-Galp-(1→4)-β-D-GlcpNAc-(1→2)-α -D-Manp-(1→O)(CH2)7CH3 mimics to explore the substrate specificity of sialyltransferases and trans-sialidases

DOI: 10.1002/ejoc.200300293

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