Convergent synthesis of the tetrasaccharide repeating unit of the cell wall lipopolysaccharide of Escherichia coli O40

Article abstract of DOI:10.3762/bjoc.8.230

A tetrasaccharide repeating unit corresponding to the cell-wall lipopolysaccharide of E. coli O40 was synthesized by using a convergent block glycosylation strategy. A disaccharide donor was coupled to a disaccharide acceptor by a stereoselective glycosyl

Full text of DOI:10.3762/bjoc.8.230

Convergent synthesis of the tetrasaccharide
repeating unit of the cell wall lipopolysaccharide
of Escherichia coli O40
Abhijit Sau and Anup Kumar Misra*
Full Research Paper
Open Access
Address:
Beilstein J. Org. Chem. 2012, 8, 2053–2059.
Bose Institute, Division of Molecular Medicine, P-1/12, C.I.T. Scheme
VII-M, Kolkata-700054, India; FAX: +91-33-2355 3886
doi:10.3762/bjoc.8.230
Received: 30 August 2012
Accepted: 01 November 2012
Published: 22 November 2012
Email:
Anup Kumar Misra* - akmisra69@gmail.com
* Corresponding author
Associate Editor: S. Flitsch
Keywords:
© 2012 Sau and Misra; licensee Beilstein-Institut.
License and terms: see end of document.
Escherichia coli; glycosylation; lipopolysaccharide; O-antigen;
tetrasaccharide
Abstract
A tetrasaccharide repeating unit corresponding to the cell-wall lipopolysaccharide of E. coli O40 was synthesized by using a
convergent block glycosylation strategy. A disaccharide donor was coupled to a disaccharide acceptor by a stereoselective glycosyl-
ation. A 2-aminoethyl linker was chosen as the anomeric protecting group at the reducing end of the tetrasaccharide. All glycosyl-
ation steps are significantly high yielding and stereoselective.
Introduction
Infantile diarrhoea is one of the major causes of morbidity and and capsular antigens [6]. Diarrhoea-causing E. coli strains are
mortality in infancy in developing countries [1]. Among several broadly classified in four categories: (a) Enteropathogenic
factors, Escherichia coli (E. coli) infection is one of the major E. coli infects through the production of heat-labile and heat-
causes of diarrhoeal disease in the developing countries [2]. stable toxins; (b) enteroinvasive E. coli acts through the inva-
E. coli are Gram-negative opportunistic pathogens and belong sion of the host body; (c) enteropathogenic E. coli infects by
to the genus Enterobacteriaceae. In general, E. coli is consid- adhering to the membrane of the host intestine; and (d) vero-
ered as a friendly organism present in the normal intestinal flora toxin E. coli infects by the production of verotoxin or shiga
of humans and animals and can kill harmful bacteria by toxin [7]. Recently, Zhao et al. reported the structure of the
producing vitamins and other immunostimulants [3]. However, repeating unit of the cell-wall antigenic lipopolysaccharide of
a number of E. coli strains acquire virulence factors and cause E. coli O40 [8], which contains two D-galactosyl moieties with
severe intestinal and urinary-tract infections [4,5]. E. coli alpha and beta linkage, one beta-linked D-glucosamine and one
serotypes are generally classified based on the somatic, flagella beta-linked D-mannosyl moiety (Figure 1).
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Beilstein J. Org. Chem. 2012, 8, 2053–2059.
Figure 1: Structure of the tetrasaccharide repeating unit of the cell-wall lipopolysaccharide of Escherichia coli O40.
Although several therapeutics have appeared in the past to concept for the orthogonal activation of thioglycoside during
control the diarrheal epidemics caused by E. coli infections, the synthesis of disaccharide derivative 9 [19]; (d) use of
emergence of resistant strains is a serious concern in the devel- aminoethyl linker as the anomeric protecting group; (e) removal
opment of therapeutics against this organism. Since, bacterial of benzyl groups using a combination of triethylsilane and
cell-wall lipopolysaccharides play important roles in the patho- Pd(OH)2–C [20]; and (f) preparation of β-D-mannosidic moiety
genicity of the virulent strains, it would be pertinent to develop from the β-D-glucoside [13].
glycoconjugate therapeutics based on the cell-wall oligosaccha-
ride haptens to reduce the number of infections [9-12]. In order Benzylation of 2-azidoethyl 3-O-benzyl-4,6-O-benzylidene-β-
to evaluate the therapeutic efficacy of the glycoconjugate D-mannopyranoside (2) [13] (prepared from D-glucose in nine
derivatives it is essential to have a significant quantity of oligo- steps) by using benzyl bromide and sodium hydroxide [21] fol-
saccharides, which is difficult to isolate from natural sources. lowed by reductive ring opening of the 4,6-O-benzylidene
Therefore, the development of a chemical synthetic strategy for acetal with triethylsilane and iodine [22] furnished compound 6
the synthesis of the oligosaccharides and their close analogues in 82% yield. Stereoselective glycosylation of compound 6 with
can add momentum towards the preparation of glycoconjugate- thioglycoside derivative 3 in the presence of a combination of
based therapeutics. In this perspective, we report herein a N-iodosuccinimide (NIS) and HClO4–SiO2 [17] gave disaccha-
concise chemical synthesis of the tetrasaccharide repeating unit ride derivative 7 in a 77% yield. Formation of compound 7 was
of the cell-wall lipopolysaccharide of E. coli O40, using a confirmed from its spectral analysis [signals at δ 4.73 (d, J =
convergent block synthetic strategy.
8.0 Hz, H-1B), 4.41 (br s, H-1A) in the 1H NMR and at δ 101.6
(C-1A), 100.7 (C-1B) in the 13C NMR spectra respectively].
Saponification of compound 7 by using sodium methoxide fol-
Results and Discussion
The target tetrasaccharide 1 as its 2-aminoethyl glycoside was lowed by 3,4-O-isopropylidenation with 2,2-dimethoxypropane
synthesized by a stereoselective glycosylation of a disaccharide and p-toluenesulfonic acid [23] furnished disaccharide deriva-
acceptor 8 and a disaccharide thioglycoside donor 9 using a tive 8 in 74% yield (Scheme 1).
[2 + 2] block synthetic strategy. The disaccharide intermediates
were synthesized from the suitably protected monosaccharide In a separate experiment, stereoselective glycosylation of thio-
derivatives 2 [13], 3 [14], 4 [15] and 5 [16], which were glycoside derivative 4 with the thioglycoside acceptor 5 in the
prepared from the commercially available reducing sugars, by presence of a combination of NIS and HClO4–SiO2 [17] in
applying a series of functional group protection–deprotection dichloromethane–diethyl ether furnished disaccharide thiogly-
methodologies (Figure 2). The synthetic strategy has a number coside derivative 9 in a 74% yield together with a minor quan-
of notable features, which include (a) stereoselective [2 + 2] tity of its other isomer (≈5%), which was separated by column
block glycosylation; (b) application of general glycosylation chromatography. Formation of compound 9 was confirmed
reactions by using thioglycosides as glycosyl donors and a from its spectral analysis [δ 5.51 (d, J = 3.5 Hz, H-1D), 5.37 (d,
combination of N-iodosuccinimide (NIS) and perchloric acid J = 10.5 Hz, H-1C) in the 1H NMR and δ 97.4 (C-1D), 83.0
supported over silica (HClO4–SiO2) [17,18] as glycosyl acti- (C-1C) in the 13C NMR spectra, respectively]. During the syn-
vator; (c) exploitation of the armed–disarmed glycosylation thesis of compound 9, thioglycoside 4 acted as glycosyl donor
Figure 2: Structure of the synthesized tetrasaccharide 1 and its synthetic precursors.
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Scheme 1: Synthesis of disaccharide derivative 8. Reagents and conditions: (a) benzyl bromide, NaOH, DMF, room temperature, 1 h; (b) Et3SiH, I2,
CH3CN, 0 °C, 1 h, 82%; (c) NIS, HClO4–SiO2, MS 4Å, CH2Cl2, −25 °C, 1 h, 77%; (d) 0.1 M CH3ONa, CH3OH, room temperature, 3 h; (e) 2,2-
dimethoxypropane, p-TsOH, DMF, room temperature, 5 h, 74%.
and thioglycoside 5 acted as orthogonal glycosyl acceptor [20] to furnish target compound 1, which was purified through a
because of the difference in their reactivity following the Sephadex® LH-20 column to give pure compound 1 in 60%
“armed–disarmed glycosylation” concept [19,24] (Scheme 2). overall yield. Spectral data of compound 1 confirmed its forma-
tion [signals at δ 5.31 (d, J = 8.5 Hz, H-1C), 5.15 (d, J = 3.5 Hz,
H-1D), 4.63 (br s, H-1A), 4.34 (d, J = 8.5 Hz, H-1B) in the
1H NMR and at δ 100.7 (C-1B), 100.6 (C-1B), 99.6 (2 C, C-1A,
C-1C) in the 13C NMR] (Scheme 3).
Conclusion
In summary, synthesis of a tetrasaccharide repeating unit
corresponding to the cell-wall lipopolysaccharide of E. coli O40
was achieved by using a convergent [2 + 2] block synthetic
strategy. The yields are excellent in all reactions. A general
reaction condition was used in all glycosylation reactions. All
intermediates and final compounds were characterized by their
spectral analysis. The armed–disarmed glycosylation concept
was applied for the synthesis of disaccharide derivative 9. A
2-Aminoethyl linker was used as the anomeric protecting group.
Scheme 2: Synthesis of disaccharide derivative 9. Reagents and
conditions: (a) NIS, HClO4–SiO2, MS 4Å, CH2Cl2–Et2O, −25 °C, 1 h,
74%.
Iodonium ion promoted [2 + 2] stereoselective glycosylation of
compound 8 and compound 9 in the presence of NIS and
HClO4–SiO2 [17] furnished tetrasaccharide derivative 10 in
71% yield. Formation of compound 10 was confirmed by its
spectral analysis [signals at δ 101.6 (C-1B), 101.5 (PhCH),
100.8 (C-1C), 100.2 (C-1A), 97.3 (C-1D) in the 13C NMR spec-
trum]. Compound 10 was subjected to a sequence of reactions
involving (a) removal of N-phthalimido group by using
hydrazine hydrate [25]; (b) N-acetylation by using acetic anhy-
dride and pyridine; (c) removal of isopropylidene ketal and
benzylidene acetal by acid hydrolysis; and finally (d) removal
of benzyl ethers by using triethylsilane and 20% Pd(OH)2–C
Experimental
General methods: All reactions were monitored by thin-layer
chromatography over silica-gel-coated TLC plates. The spots on
TLC were visualized by warming ceric sulfate (2% Ce(SO4)2 in
2 N H2SO4)-sprayed plates on a hot plate. Silica gel 230–400
mesh was used for column chromatography. 1H and 13C NMR
spectra were recorded on Brucker Avance 500 MHz by using
Scheme 3: Synthesis of target tetrasaccharide 1. Reagents and conditions: (a) NIS, HClO4–SiO2, CH2Cl2, −15 °C, 1 h, 71%; (b) NH2NH2·H2O, EtOH,
90 °C, 5 h; (c) acetic anhydride, pyridine, room temperature, 2 h; (d) 80% aq AcOH, 80 °C, 1.5 h; (e) triethylsilane, 20% Pd(OH)2–C, CH3OH, 6 h,
room temperature, overall 60%.
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Beilstein J. Org. Chem. 2012, 8, 2053–2059.
CDCl3 as solvent and TMS as internal reference, unless stated argon. The reaction mixture was cooled to −25 °C, and
otherwise. Chemical shift values are expressed in δ ppm. N-iodosuccinimide (NIS; 0.8 g, 3.55 mmol) and HClO4–SiO2
MALDI-MS were recorded on a Bruker Daltronics mass spec- (25.0 mg) were added to it. After being stirred at same tempera-
trometer. Commercially available grades of organic solvents ture for 1 h the reaction mixture was filtered through a Celite®
of adequate purity were used in all reactions. HClO4–SiO2 bed and washed with CH2Cl2 (100 mL). The organic layer was
was prepared following the method reported in the literature successively washed with 5% Na2S2O3, saturated NaHCO3 and
[18].
water, and then dried (Na2SO4) and concentrated under reduced
pressure to give the crude product. The crude product was puri-
2-Azidoethyl 2,3,6-tri-O-benzyl-β-D-mannopyranoside (6): fied over SiO2 by using hexane–EtOAc (7:1) as eluant to give
To a solution of compound 2 (2.0 g, 4.68 mmol) in dry DMF pure compound 7 (2.0 g, 77%). Yellow oil; [α]D25 −13 (c 1.0,
(10 mL) were added benzyl bromide (1.2 mL, 10.09 mmol) and CHCl3); IR (neat): 3087, 2956, 2153, 1605, 1487, 1345, 1254,
powdered NaOH (750.0 mg, 18.75 mmol) and the reaction mix- 1183, 1142, 1045, 999, 774, 734, 647, 542 cm−1; 1H NMR (500
ture was stirred at room temperature for 1 h. The reaction mix- MHz, CDCl3) δ 7.35–7.16 (m, 20H, Ar-H), 5.33 (d, J = 3.0 Hz,
ture was diluted with water (100 mL) and extracted with 1H, H-4B), 5.06 (t, J = 8.0 Hz each, 1H, H-2B), 4.87 (d, J = 12.0
CH2Cl2 (100 mL). The organic layer was washed with H2O, Hz, 1H, PhCH2), 4.84 (dd, J = 10.5, 3.5 Hz, 1H, H-3B), 4.73 (d,
dried (Na2SO4) and concentrated. The crude product was J = 8.0 Hz, 1H, H-1B), 4.72–4.71 (2 d, J = 12.0 Hz each, 2H,
passed through a short pad of SiO2 by using hexane–EtOAc PhCH2), 4.59 (d, J = 12.0 Hz, 1H, PhCH2), 4.47, 4.45 (2 d, J =
(5:1) as eluant to give the O-benzylated product (2.2 g, 91%). A 12.0 Hz each, 2H, PhCH2), 4.41 (br s, 1H, H-1A), 4.39 (d, J =
solution of the O-benzylated product (2.2 g, 4.25 mmol) in dry 12.0 Hz, 1H, PhCH2), 4.19 (d, J = 12.0 Hz, 1H, PhCH2), 4.14
CH3CN (20 mL) was cooled to 0 °C. To the cooled reaction (t, J = 9.0 Hz each, 1H, H-4A), 4.10–4.05 (m, 1H, OCH2), 3.92
mixture were added Et3SiH (1.4 mL, 8.76 mmol) and I2 (d, J = 3.0 Hz, 1H, H-2A), 3.76–3.70 (m, 2H, H-6abA),
(250.0 mg, 0.98 mmol), and the reaction mixture was stirred at 3.65–3.60 (m, 1H, OCH2), 3.54–3.49 (m, 2H, H-5B, CH2N3),
the same temperature for 1 h. The reaction mixture was diluted 3.47 (dd, J = 10.0, 3.0 Hz, 1H, H-3A), 3.42–3.38 (m, 1H, H-5A),
with CH2Cl2 (100 mL) and the organic layer was successively 3.32–3.22 (m, 3H, H-6abB, CH2N3), 1.99, 1.95, 1.90 (3 s, 9H, 3
washed with saturated NaHCO3 and H2O, and then dried COCH3); 13C NMR (125 MHz, CDCl3) δ 169.9, 169.8, 169.4
(Na2SO4) and concentrated. The crude product was purified (3 COCH3), 138.5–126.8 (Ar-C), 101.6 (C-1A), 100.7 (C-1B),
over SiO2 by using hexane–EtOAc (4:1) as eluant to give pure 80.6 (C-5B), 75.7 (C-5A), 74.7 (C-2A), 74.3 (2 C, C-4A,
compound 6 (1.7 g, overall 82%). White solid; mp 89–90 °C; PhCH2), 73.7 (PhCH2), 73.3 (PhCH2), 71.7 (C-5B), 71.5
[α]D25 −97 (c 1.0, CHCl3); IR (KBr): 3293, 2845, 2110, 1497, (PhCH2), 71.3 (C-3B), 70.0 (C-2B), 68.6 (C-6A), 68.5 (OCH2),
1454, 1365, 1310, 1119, 1065, 779, 659, 599 cm−1; 1H NMR 67.4 (C-4B), 66.9 (C-6B), 50.8 (CH2N3), 20.7, 20.6, 20.5
(500 MHz, CDCl3) δ 7.43–7.23 (m, 15H, Ar-H), 4.98 (d, J = (COCH3); MALDI-MS: 920.3 [M + Na]+; Anal. calcd for
12.5 Hz, 1H, PhCH2), 4.75 (d, J = 12.5 Hz, 1H, PhCH2), 4.61, C48H55N3O14: C, 64.20; H, 6.17; found: C, 64.06; H, 6.35.
4.58 (2 d, J = 12.0 Hz, 2H, PhCH2), 4.48 (br s, 1H, H-1), 4.45
(d, J = 12.0 Hz, 1H, PhCH2), 4.32 (d, J = 12.0 Hz, 1H, PhCH2), 2-Azidoethyl O-(6-O-benzyl-3,4-O-isopropylidene-β-D-
4.15–4.11 (m, 1H,-OCH2-), 3.96 (br s, 1H, H-2), 3.94 (t, J = 9.5 galactopyranosyl)-(1→4)-2,3,6-tri-O-benzyl-β-D-manno-
Hz each, 1H, H-4), 3.85 (dd, J = 10.5, 3.5 Hz, 1H, H-6a), 3.75 pyranoside (8): A solution of compound 7 (1.8 g, 2.0 mmol) in
(dd, J = 10.5, 6.5 Hz, 1H, H-6b), 3.66–3.62 (m, 1H, OCH2-), 0.1 M CH3ONa (25 mL) was stirred at room temperature for
3.58–3.53 (m, 1H, CH2N3), 3.46–3.42 (m, 1H, H-5), 3.34–3.30 2 h. The reaction mixture was neutralized with Dowex 50W X8
(m, 1H, CH2N3), 3.29 (dd, J = 10.0, 3.5 Hz, 1H, H-3); (H+) resin, filtered and concentrated. To a solution of the de-O-
13C NMR (125 MHz, CDCl3) δ 138.6–127.4 (Ar-C), 101.8 acetylated product in dry DMF (10 mL) was added 2,2-
(C-1), 81.3 (C-3), 75.4 (C-5), 74.4 (PhCH2), 73.7 (PhCH2), 73.5 dimethoxypropane (0.7 mL, 5.69 mmol) followed by p-TsOH
(C-5), 71.1 (PhCH2), 70.7 (C-6), 68.6 (OCH2), 68.0 (C-2), 50.9 (0.2 g) and the reaction mixture was stirred at room tempera-
(CH2N3); ESI-MS: 542.2 [M + Na]+; Anal. calcd for ture for 5 h. The reaction was quenched with Et3N (1 mL), the
C29H33N3O6: C, 67.04; H, 6.40; found: C, 66.90; H, 6.58. solvents were removed under reduced pressure, and the crude
reaction mixture was diluted with CH2Cl2 (100 mL). The
2-Azidoethyl O-(2,3,4-tri-O-acetyl-6-O-benzyl-β-D- organic layer was washed with saturated NaHCO3, dried
galactopyranosyl)-(1→4)-2,3,6-tri-O-benzyl-β-D-manno- (Na2SO4) and concentrated to give the crude product, which
pyranoside (7): To a solution of compound 3 (1.4 g, was purified over SiO2 by using hexane–EtOAc (2:1) as eluant
3.18 mmol) and compound 6 (1.5 g, 2.88 mmol) in anhydrous to give pure compound 8 (1.2 g, 74%). Yellow oil; [α]D25 −21
CH2Cl2 (10 mL) was added MS 4Å (2.0 g), and the reaction (c 1.0, CHCl3); IR (neat): 3418, 3030, 2926, 2198, 1743, 1711,
mixture was stirred at room temperature for 30 min under 1646, 1390, 1253, 1099, 1053, 864, 754, 667, 531 cm−1;
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Beilstein J. Org. Chem. 2012, 8, 2053–2059.
1H NMR (500 MHz, CDCl3) δ 7.30–7.14 (m, 20H, Ar-H), 4.84 83.0 (C-1C), 81.7 (C-4D), 78.1 (C-3D), 75.4 (C-2D), 74.8
(d, J = 12.5 Hz, 1H, PhCH2), 4.63–4.40 (m, 6H, PhCH2), 4.38 (C-5D), 74.7 (PhCH2), 73.3 (2 C, C-3C, PhCH2), 72.8 (PhCH2),
(d, J = 8.0 Hz, 1H, H-1B), 4.33 (br s, 1H, H-1A), 4.30 (d, J = 71.8 (PhCH2), 70.1 (C-5C), 69.4 (C-4C), 68.8 (C-6D), 67.7
12.5 Hz, 1H, PhCH2), 4.22 (t, J = 9.5 Hz each, 1H, H-4A), (C-6C), 54.2 (C-2C), 24.0 (SCH2CH3), 14.9 (SCH2CH3);
4.02–3.98 (m, 2H, H-2A, OCH2), 3.91 (dd, J = 10.0, 3.5 Hz, MALDI-MS: 986.3 [M + Na]+; Anal. calcd for C57H57NO11S:
H-3B), 3.83 (dd, J = 12.0, 5.5 Hz, 1H, H-6aB), 3.81 (d, J = 2.0 C, 71.01; H, 5.96; found: C, 70.88; H, 6.13.
Hz, 1H, H-4B), 3.76 (dd, J = 12.0, 2.0, Hz, 1H, H-6bB), 3.71 (br
s, 1H, H-5B), 3.70–3.68 (m, 1H, OCH2), 3.62–3.58 (m, 1H, 2 - A z i d o e t h y l O - ( 2 , 3 , 4 , 6 - t e t r a - O - b e n z y l - α - D -
H-6aA), 3.56–3.51 (m, 1H, CH2N3), 3.49 (dd, J = 8.0 Hz each, galactopyranosyl)-(1→3)-O-(4,6-O-benzylidene-2-deoxy-2-
1H, H-2B), 3.47–3.43 (m, 1H, H-6bA), 3.42–3.37 (m, 2H, H-3A, N-phthalimido-β-D-glucopyranosyl)-(1→2)-O-(6-O-benzyl-
H-5A), 3.25–3.19 (m, 1H, CH2N3), 1.42, 1.25 (2 s, 6H, 2 3,4-O-isopropylidene-β-D-galactopyranosyl)-(1→4)-2,3,6-
C(CH3)3); 13C NMR (125 MHz, CDCl3) δ 138.7–127.3 (Ar-C), tri-O-benzyl-β-D-mannopyranoside (10): To a solution of
109.8 (C(CH3)2), 102.5 (C-1B), 101.9 (C-1A), 80.6 (C-2B), 78.9 compound 8 (1.0 g, 1.23 mmol) and compound 9 (1.3 g,
(C-3B), 75.2 (C-5A), 74.3 (PhCH2), 74.2 (C-3A), 74.0 (C-4B), 1.35 mmol) in anhydrous CH2Cl2 (10 mL) was added MS 4Å
73.8 (C-2A), 73.5 (PhCH2), 73.4 (2C, C-4A, PhCH2), 72.4 (2.0 g), and reaction mixture was stirred at room temperature
(C-5B), 71.3 (PhCH2), 69.4 (OCH2), 69.3 (C-6B), 68.5 (C-6A), for 30 min under argon. The reaction mixture was cooled to
50.8 (CH2N3), 28.2, 26.4 (C(CH3)2); MALDI-MS: 834.3 [M + −25 °C and NIS (350.0 mg, 1.55 mmol) and HClO4–SiO2
Na]+; Anal. calcd for C45H53N3O11: C, 66.57; H, 6.58; found: (10.0 mg) were added to it. After being stirred at same tempera-
C, 66.42; H, 6.75.
ture for 1 h the reaction mixture was filtered through a Celite®
bed and washed with CH2Cl2 (100 mL). The organic layer was
Ethyl O-(2,3,4,6-tetra-O-benzyl-α-D-galactopyranosyl)- successively washed with 5% Na2S2O3, saturated NaHCO3 and
(1→3)-4,6-O-benzylidene-2-deoxy-2-N-phthalimido-1-thio- water, and then dried (Na2SO4) and concentrated under reduced
β-D-glucopyranoside (9): To a solution of compound 4 (1.4 g, pressure to give the crude product. The crude product was puri-
2.39 mmol) and compound 5 (1.0 g, 2.26 mmol) in anhydrous fied over SiO2 by using hexane–EtOAc (7:1) as eluant to give
CH2Cl2–Et2O (10 mL; 1:1 v/v) was added MS 4Å (2.0 g), and pure compound 10 (1.5 g, 71%). White solid; mp 65–66 °C;
reaction mixture was stirred at room temperature for 30 min [α]D25 +34 (c 1.0, CHCl3); IR (KBr): 3423, 3063, 3030, 2871,
under argon. The reaction mixture was cooled to −25 °C and 2105, 1776, 1744, 1715, 1497, 1454, 1389, 1239, 1102, 1060,
NIS (550.0 mg, 2.44 mmol) and HClO4–SiO2 (15.0 mg) were 874, 737, 721, 697, 600, 530 cm−1; 1H NMR (500 MHz,
added. After being stirred at same temperature for 1 h the reac- CDCl3) δ 7.76–6.96 (m, 49H, Ar-H), 5.52 (d, J = 3.5 Hz, 1H,
tion mixture was filtered through a Celite® bed and washed H-1D), 5.42 (d, J = 8.0 Hz, 1H, H-1C), 5.25 (s, 1H, PhCH),
with CH2Cl2 (100 mL). The organic layer was successively 4.86–4.45 (m, 13H, PhCH2), 4.41 (d, J = 9.5 Hz, 1H, H-1B),
washed with 5% Na2S2O3, saturated NaHCO3 and water, and 4.37 (t, J = 8.0 Hz, 1H, H-2C), 4.29 (d, J = 11.5 Hz, 1H,
then dried (Na2SO4) and concentrated under reduced pressure PhCH2), 4.24–4.20 (m, 4H, H-1A, H-3C, H-4A, OCH2),
to give the crude product. The crude product was purified over 4.13–4.08 (m, 2H, H-2A, OCH2), 3.91 (br s, 2H, PhCH2),
SiO2 by using hexane–EtOAc (7:1) as eluant to give pure com- 3.89–3.85 (m, 3H, H-2D, H-3B, H-3D), 3.84–3.75 (m, 3H, H-2B,
pound 9 (1.6 g, 74%). White solid; mp 67–68 °C; [α]D25 +39 (c H-4C, H-4D), 3.70 (t, J = 10.5 Hz each, 1H, H-6aB), 3.64–3.60
1.0, CHCl3); IR (KBr): 3417, 3063, 2870, 1774, 1715, 1610, (m, 4H, H-4B, H-5B, H-5D, H-6bB), 3.58–3.43 (m, 5H, H-3A,
1495, 1485, 1385, 1216, 1099, 1023, 914, 753, 719 cm−1; H-6aA, H-6aC, H-6abD), 3.42–3.36 (m, 4H, H-5A, H-6bA, H-6bC,
1H NMR (500 MHz, CDCl3) δ 7.76–6.91 (m, 29H, Ar-H), 5.51 CH2N3), 3.34–3.26 (m, 2H, H-5C, CH2N3), 1.27, 1.25 (2 s, 6H,
(d, J = 3.5 Hz, 1H, H-1D), 5.37 (d, J = 10.5 Hz, 1H, H-1C), 5.32 2 CH3); 13C NMR (125 MHz, CDCl3) δ 138.6–126.3 (Ar-C),
(s, 1H, PhCH), 4.84 (t, J = 9.5 Hz each, 1H, H-3C), 4.77–4.58 109.5 (C(CH3)2), 101.6 (C-1B), 101.5 (PhCH), 100.8 (C-1C),
(3 d, J = 12.0 Hz each, 3H, PhCH2), 4.46 (t, J = 9.5 Hz each, 100.2 (C-1A), 97.3 (C-1D), 82.9 (C-4C), 82.7 (C-5A), 80.1
1H, H-2C), 4.44 (d, J = 11.5 Hz, 1H, PhCH2), 4.34 (d, J = 11.5 (C-5c), 78.7 (C-3A), 78.1 (C-4D), 76.2 (C-2D), 75.5 (C-2B),
Hz, 1H, PhCH2), 4.29 (t, J = 9.5 Hz each, 1H, H-4C), 4.19 (d, J 74.9 (C-3B), 74.8 (C-5D), 74.7 (PhCH2), 74.6 (C-3D), 73.9
= 11.5 Hz, 1H, PhCH2), 3.86 (br s, 2H, PhCH2), 3.85 (br s, 1H, (PhCH2) 73.5 (C-3C), 73.4 (2 C, 2 PhCH2), 73.2 (PhCH2), 72.8
H-4D), 3.81 (dd, J = 10.5, 3.0 Hz, 1H, H-2D), 3.72–3.67 (m, 3H, (PhCH2), 72.6 (C-2A), 72.0 (PhCH2), 71.7 (2 C, C-4B, PhCH2),
H-3D, H-5C, H-6aD), 3.58 (br s, 1H, H-5D), 3.33–3.31 (m, 1H, 69.3 (C-4A), 68.9 (C-6C), 68.8 (OCH2), 68.7 (C-6A), 68.4
H-6bD), 3.23–3.19 (m, 1H, H-6aC), 2.80–2.77 (m, 1H, H-6bC), (OCH2), 67.7 (2 C, C-6B, C-6D), 65.4 (C-5B), 56.0 (C-2C), 50.8
2.67–2.56 (m, 2H, SCH2CH3), 1.12 (t, J = 7.5 Hz each, 3H, (CH2N3), 27.6, 25.7 (C(CH3)2); MALDI-MS: 1735.7 [M +
SCH2CH3); 13C NMR (125 MHz, CDCl3) δ 168.1, 167.9 Na]+; Anal. calcd for C100H104N4O22: C, 70.08; H, 6.12; found:
(PhthCO), 138.9–123.1 (Ar-C), 101.7 (PhCH), 97.4 (C-1D), C, 69.94; H, 6.30.
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References
2-Aminoethyl (α-D-galactopyranosyl)-(1→3)-(2-acetamido-
2-deoxy-β-D-glucopyranosyl)-(1→2)-(β-D-galacto-
pyranosyl)-(1→4)-β-D-mannopyranoside (1): To a solution
of compound 10 (500.0 mg, 0.29 mmol) in EtOH (5 mL) was
added NH2NH2·H2O (0.1 mL) and the reaction mixture was
stirred at 90 °C for 5 h. The solvents were removed under
reduced pressure, and a solution of the crude product in acetic
anhydride–pyridine (2 mL, 1:1 v/v) was kept at room tempera-
ture for 2 h and then concentrated. A solution of the crude prod-
uct in 80% aq AcOH (10 mL) was stirred at 80 °C for 1.5 h and
then concentrated. To a solution of the crude product in CH3OH
(5 mL) were added Et3SiH (1.5 mL, 9.39 mmol) and 20%
Pd(OH)2–C (100.0 mg) and the reaction mixture was stirred at
room temperature for 6 h. The reaction mixture was filtered
through a Celite® bed and washed with CH3OH–H2O (2:1).
The solvents were removed under reduced pressure and the
product was passed through a Sephadex® LH-20 column by
using CH3OH–H2O (3:1) as eluant to furnish pure compound 1
(135.0 mg, 60%). Glass; [α]D25 +29 (c 1.0, H2O); IR (KBr):
3436, 2948, 1619, 1369, 1162, 669 cm−1; 1H NMR (500 MHz,
D2O) δ 5.31 (d, J = 8.5 Hz, 1H, H-1C), 5.15 (d, J = 3.5 Hz, 1H,
H-1D), 4.63 (br s, 1H, H-1A), 4.48 (t, J = 10.5 Hz each, 1H,
H-3C), 4.34 (d, J = 8.5 Hz, 1H, H-1B), 4.09 (t, J = 10.0 Hz each,
1H, H-2C), 4.05–3.92 (m, 4H, H-2A, H-4D, H-5D, OCH2a),
3.90–3.80 (m, 4H, H-2B, H-3A, H-6aB, OCH2b), 3.78–3.57 (m,
11H, H-3D, H-4B, H-4C, H-6abA, H-6bB, H-6abC, H-6abD),
3.55–3.47 (m, 2H, H-2D, H-3B), 3.45–3.40 (m, 2H, H-5A,
H-5B), 3.35–3.33 (m, 1H, H-5C), 3.20–3.15 (m, 2H, CH2NH2),
2.06 (s, 3H, COCH3); 13C NMR (125 MHz, D2O) δ 171.5
(COCH3), 100.7 (C-1B), 100.6 (C-1B), 99.6 (2C, C-1A, C-1C),
81.7 (C-3B), 79.0 (C-3C), 78.5 (C-4D), 77.0 (C-3D), 75.5
(C-4A), 75.1 (C-4B), 73.5 (C-2D), 73.3 (C-5A), 71.3 (C-2A),
70.7 (C-4C), 70.6 (C-3A), 69.8 (2 C, C-5C, C-5D), 68.3 (2C,
C-2B, C-5B), 65.5 (OCH2), 60.6 (C-6B), 60.5 (C-6C), 60.2
(C-6A), 59.2 (C-6D), 55.7 (C-2C), 39.5 (CH2NH2), 23.1
(COCH3); MALDI-MS: 799.2 [M + Na]+; Anal. calcd for
C28H48N4O21: C, 43.30; H, 6.23; found: C, 43.14; H, 6.45.
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1D and 2D NMR spectra of compounds 1 and 6–10.
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Acknowledgements
A. S. thanks CSIR, New Delhi for providing a Senior Research
Fellowship. This work was supported by DST, New Delhi
(Project No. SR/S1/OC-83/2010).
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Beilstein J. Org. Chem. 2012, 8, 2053–2059.
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