International Journal of Carbohydrate Chemistry

International Journal of Carbohydrate Chemistry / 2010 / Article

Research Article | Open Access

Volume 2010 |Article ID 594247 |

Galina Pazynina, Vitaly Nasonov, Ivan Belyanchikov, Reinchard Brossmer, Maxim Maisel, Alexander Tuzikov, Nicolai Bovin, "Koenigs-Knorr Glycosylation with Neuraminic Acid Derivatives", International Journal of Carbohydrate Chemistry, vol. 2010, Article ID 594247, 8 pages, 2010.

Koenigs-Knorr Glycosylation with Neuraminic Acid Derivatives

Academic Editor: Richard D. Cummings
Received13 Jul 2010
Revised15 Sep 2010
Accepted21 Oct 2010
Published02 Dec 2010


Earlier we reported a convenient and efficient method of preparing α2-6 sialooligosaccharides in conditions of Koenigs-Knorr reaction. The use of Ag2CO3 allowed carrying out α2-6 sialylation of galacto-4,6-diol of mono- and disaccharides with chloride of acetylated N-acetylneuraminic acid methyl ester as glycosyl donor. In this study we applied this approach to other derivatives of neuraminic acid, namely, Neu5Gc, 9-deoxy-9-NAc-Neu5Ac, Neu5Acα2-8Neu5Ac, and Neu5Acα2-8Neu5Acα2-8Neu5Ac as glycosyl donors; eight compounds were synthesized: Neu5Gcα-O(CH2)3NH2 (8), Neu5Gcα2-6Galβ1-4GlcNAcβ-O(CH2)3NH2 (10), 9-deoxy-9-NAc-Neu5Ac-O(CH2)3NH2 (15), 9-deoxy-9-NAc-Neu5Acα2-6Galβ1-4GlcNAcβ-O(CH2)3NH2 (17), Neu5Acα2-8Neu5Acα-O(CH2)3NH2(23) Neu5Acα2-8Neu5Acα-OCH3 (24), Neu5Acα2-8Neu5Acα-OCH2(p-C6H4)NHCOCH2NH2 (25), and Neu5Acα2-8Neu5Acα2-8Neu5Acα-O(CH2)3NH2 (32). These sialosides were used for characterization of siglecs and other carbohydrate-binding proteins.

1. Introduction

Sialic acid family comprises over 40 neuraminic acid versions and monosaccharide derivatives; the number of biologically relevant sialic acid oligosaccharides is also very high. Sialic acids are a part of the most important molecules of life, since they occupy the terminal position on cell membrane glycoproteins and glycolipids. Given their location and ubiquitous distribution, sialic acids can mediate or modulate a wide range of physiological and pathological processes and play a role not only in the protection and adaptation of life, but also in being utilized by life-threatening infectious microorganisms [1]. Several families of sialic acid-binding proteins have been discovered over the last few decades, including mammalian selectins and siglecs [2]. Only a limited number of sialomolecules are available as the tools for the study of mammalian and microbial lectins and sialo-modifying enzymes. Due to this, the search of simple and efficient synthetic ways for sialosaccharides is very relevant. The methods of sialylation described in the literature [35] rarely give high yields and α-stereoselectivity but suggest complex modifications of donors; introduction and removal of substituents at C-3, C-4, C-5, and C-9 in order to improve sialylation stereoselectivity presume complex multistage synthesis of such compounds. In case of the use of 2-xantates, thiophosphates and various imidates, sialylation usually proceeds at −50–70°C and requires strong Lewis acids as promoters, this posing the corresponding limitations to the repertoire of protective groups in glycosyl acceptor. Recently we have described a simple and efficient method of the synthesis of α-2-6 sialooligosaccharides in conditions of Koenigs-Knorr reaction [6] when the simplest possible sialyl donor is used at room temperature ang gives high α-stereoselectivity. It was interesting to study the applicability of this approach to other neuraminic acid derivatives (Neu5Gc, 9-deoxy-9-NAc-Neu5Ac, Neu5Acα2-8Neu5Ac, and Neu5Acα2-8Neu5Acα2-8Neu5Ac). Here, we describe practical chemical syntheses of several spacer-armed sialic acid glycosides, with especial interest to N-glycolyl-neuraminic acid version playing role in cancer immunology and transplantation, and also α2-8 oligosaccharides known as key motif of polysialic acids, numerous gangliosides, and serving as affinity ligand for siglec-7 [7].

Peracetylated methyl esters of Neu5Gc (6) [8], 9-deoxy-9-NAc-Neu5Ac (13), Neu5Acα2-8Neu5Ac (19 and 20) [9], and Neu5Acα2-8Neu5Acα2-8Neu5Ac (2730) [10] were synthesized starting from free saccharides by esterification in acidic conditions (MeOH/HCl) with subsequent acetylation (Ac2O/pyridine).

2-Chloroderivatives 7, 14, , and 31, Sug-Cl, were synthesized from the corresponding peracetylated methyl esters by treatment with HCl in CHCl3 for 1.5–2 days and were used without further purification.

Glycosylation was performed in dry CH2Cl2 in the presence of Ag2CO3 or Ag2CO3/AgOTf (silver trifluoromethanesulfonate) and molecular sieves 4Å (MS 4Å) at room temperature for 1–7 days; the reaction conditions and yields are given in the Table 1. In this study we have used equivalent amounts of glycosyl donor (Sug-Cl) and acceptor or excess of acceptor. The corresponding glycals were found to be the main side products in all glycosylation reactions. Deprotection resulted in the mixture of the target product with glycal and unreacted acceptor, easily separated using cation-exchange chromatography on Dowex H+ resin [11]: elution with water gave glycal (acid), whereas the product (amino acid) had been retained on the column and was completely eluted with 1 M aqueous pyridine; unreacted glycosyl acceptor (amine) was eluted with 1 M NH4OH. In the case of disialic and trisialic glycosides partial lactonization occurred during purification on Dowex H+, so the lactone was treated with aqueous NaOH to give corresponding acid.


+β, %

α, %
α/β b

1Neu5Ac (3)HO(CH2)3NHCOCF3522.6 (62 h)  47171  : 2 [8]
HO(CH2)3NHCOCF3230.10.2 (5 h)473695 : 5

3Neu5Gc (7)HO(CH2)3NHCOCF33338685792 : 8
130.1710301243 : 57

Neu5Ac (14)
HO(CH2)3NHCOCF313315n.d.29Traces of β
16517n.d.25Traces of β

7Neu5Ac2-8HO(CH2)3NHCOCF3237  2320not done94 : 6
8Neu5Ac 2-8HO(CH2)3NHCOCF3230.11363290 : 10
9(21+22)MeOH10060.15324n.d.40Traces of β
10HOCH2(p-C6H4)NHCOCH2NHBoc25 3  25201050 : 50
11HOCH2(p-C6H4)NHCOCH2NHBoc460.63321239 : 61

Neu5Ac2-8 (31)
HO(CH2)3NHCOCF333 7  32332781 : 19
13HO(CH2)3NHCOCF3330.5737not done67 : 33

Preparative yield of unprotected glycoside Sug-OR calculated on protected precursor of Sug-Cl (n.d.: not determined);
/β ratio according to NMR data;
series of experiments demonstrated that these conditions are optimal for the reaction in the presence of AgOTf;
Recovery of the acceptor in the form LacNAcβ-O(CH2)3NH2 (11) 58%;
Recovery of the acceptor in the form LacNAcβ-O(CH2)3NH2 (11) 66%;
Reaction did not proceed without AgOTf.

All Sug-Cl (see Table 1) have demonstrated considerably less activity in glycosylation compared to chloride of acetylated N-acetylneuraminic acid methyl ester (3) [6, 8]; in several cases Ag2CO3 promoted glycosylation (without AgOTf) either proceeded very slowly or did not proceed at all. The optimal of AgOTf added to Ag2CO3 ratio (in respect to the reaction duration and acceptable α/β anomer ratio) was found to be 10% mol using chloride of Neu5Ac (3) as glycosyl donor and HO(CH2)3NHCOCF3 as acceptor; larger amount gave more β-isomer and glycal; preferable acceptor/donor ratio was not less than 2. Glycosylation at −10°C did not improve the α/β ratio compared to room temperature.

Glycosylation with chloride of Neu5Gc (7) catalyzed by Ag2CO3 in absence of AgOTf proceeded slowly; 3-trifluoroacetamidopropanol (Scheme 1, no. 3 in Table 1) gave the mixture of anomers in 68% yield (α/β 92 : 8). Deprotected product was isolated on Dowex H+ resin (elution with 1 M Py) with additional HPLC purification; the corresponding α-glycoside (8) was isolated in 57% yield, that was better result compared to NIS/TFS-OH promoted glycosylation with Neu5GcβSEt [8].


Glycosylation of 4′,6′-diol of N-acetyllactosamine (9) with chloride of Neu5Gc 7 in presence of Ag2CO3/AgOTf (Scheme 1, no. 4 in Table 1) gave 30% of α/β mixture (43 : 57); the pure α-glycoside 10 was isolated (HPLC) in 12% yield. Unreacted acceptor 9 (58%) was recovered in deprotected form 11.

Glycosylation with chloride of 9-deoxy-9-NAc-Neu5Ac 14 in presence of only Ag2CO3 proceeded similarly to the reaction with Neu5Ac (Scheme 2, no. 5 and no. 6 in Table 1). 3-Trifluoroacetamidopropanol and 4′,6′-diol 16 gave comparable yields of corresponding α-glycosides (29% of 15 and 25% of 17); β anomers were observed in trace amounts. Unreacted acceptor 16 was recovered (66%) in deprotected form 11.


Methylation followed by acetylation of Neu5Acα2-8Neu5Ac (18) or Neu5Acα2-8Neu5Acα2-8Neu5Ac (26) gave the mixtures of acetates of methyl esters and corresponding lactones (preparation of the lactones described in [9, 10]); the mixtures were used without separation. Acetates of disialic acid were the mixture of 19 and 20 (yield 82%); 2-Chloroderivatives used as glycosyl donor were the mixture of 21 and 22 (Scheme 3). Acetates of trisialic acid were the mixture of four substances (2730) (yield 72%); the mixture of corresponding four 2-Chloroderivatives used as glycosyl donor is designated as 31 (Scheme 4).


Glycosylation with Neu5Acα2-8Neu5Ac glycosyl chlorides in absence of AgOTf proceeded very slowly. Spacer alcohol HO(CH2)3NHCOCF3 gave ~20% yield of glycosylation product 23, α/β ratio 94 : 6. Glycosylation in the presence of AgOTf proceeded seven times faster resulting in α/β ratio 90 : 10 (32%) (Scheme 3, no. 7 and no. 8 in Table 1). Synthesis of disialic acid methyl glycoside was carried out at 100-fold excess of methanol (Scheme 3, no. 9 in Table 1). After deprotection and chromatography on silica gel, purification on Sephadex LH-20 and ion exchange to Na+ on Dowex the yield of target α-glycoside 24 was 40% (~33% on starting disialic acid) that is comparable to the literature data [12]; only the traces of β glycoside were observed.

In case of p-amidoglycyl-benzyl alcohol glycosylation with Neu5Acα2-8Neu5Ac the use of 0.1 mol of AgOTf per glycosyl chloride gave only 10% of 25; 0.6 ratio led to increase of conversion (yield of increased from 20% to 32%), but it was accompanied with the increase of β-anomer formation (Scheme 3, no. 10 and no. 11 in Table 1).

Trisialic chlorides 31 reacted with HO(CH2)3NHCOCF3 only in the presence of AgOTf; increase of mol/mol ratios from 0.1 to 0.5 on chloride gave 32 (~25% yield) accompanied by the increase of conversion degree and reduction of α/β ratio (Scheme 4, no. 12 and no. 13 in Table 1).

The structure of all the studied compounds was confirmed by high resolution 1H NMR data. Spectra of derivatives 4, 8, 10, and 24 coincided with those described before [8, 12, 13]. Chemical shift of H- (Neu5Ac, Neu5Gc and 9-deoxy-9-NAc-Neu5Ac residues) was 2.6-2.7 ppm for α-anomer and 2.2–2.4 ppm for β-anomer [3]. Upfield shift has been observed in spectra of 9-deoxy-9-NAc-Neu5Ac derivatives for H-9 (3.6 ppm) and H9′ (3.3 ppm) as compared to Neu5Ac (3.9 and 3.6 ppm, correspondingly) [14]. Glycosidic α(2-8) bond in disialic and trisialic acid derivatives was characterized by downfield position of H-8 and H-9 ( 4.0 ppm) as compared to Neu5Ac [12, 15]. The synthesized sialosides were used for characterization of siglecs [16] and other carbohydrate-binding proteins [17, 18].

2. Experimental

Neu5Ac was from Juelich Enzyme Products GmbH (Wiesbaden, Germany); 9-deoxy-9-NAc-Neu5Ac was obtained as described in [19]; Neu5Acα2-8Neu5Ac and Neu5Acα2-8Neu5Acα2-8Neu5Ac and Neu5Gc were from Nacalai Tesque Inc. (Kyoto, Japan).

2.1. Preparation of Peracetylated Methyl Esters of Neuraminic Acid Derivatives General procedure for 6, 13, 19  +  20, and 27–30

Neuraminic acid derivative (0.5 mmol of 5, 12, 18, or 26 as Na salt) was suspended in 25 ml of dry methanol cooled to 0°C followed by dropwise addition of AcCl (178 μl, 2.5 mmol). The mixture was stirred at room temperature for 2 h (clear solution is formed). The solution was evaporated at reduced pressure (water jet pump) and coevaporated three times with toluene. The residue was acetylated with 9 ml of Ac2O/Py (1 : 2) for 15–20 h at 40°C, coevaporated with toluene, and subjected to chromatography on silica gel (yields 70%–90%). Peracetylated derivatives of di- and trisialic acids ( and 27–30) were mixtures of acetates of methyl ester and lactones, which were used without separation.

2.2. Preparation of Neuraminic Acid Glycosyl Chlorides Derivatives (Sug-Cl) General procedure for 7, 14, 21 + 22, and 31

Peracetylated methyl ester of neuraminic acid derivative (1 mmol of 6, 13, , or 2730) was dissolved in 10 ml freshly distilled chloroform followed by addition of dry methanol (0.81 ml, 20 mmol). The solution was cooled (ice + salt) and AcCl (2.84 ml, 40 mmol) was added dropwise at cooling and stirring. After 30 min the reaction mixture was thoroughly sealed then warmed slowly and kept at room temperature for 1.5–2 days. The reaction mixture was evaporated at reduced pressure (water jet pump) and coevaporated with dry toluene several times to neutral pH value of the solution. Chlorides obtained in this way were used without further purification assuming quantitative yield of the reaction (the substances were stored at 18°C).

2.3. Glycosylation General procedure for 4, 8, 10, 15, 17, 23, 24, 25, and 32 (see Table 1 for details)

A solution of Sug-Cl (0.1 mmol) in dry CH2Cl2 (3 ml) and AgOTf (0.01–0.06 mmol) were added to a mixture of acceptor (0.1–0.4 mmol), Ag2CO3 (0.3–0.6 mmol), and MS 4Å (1 g) in dry methylene chloride (5 ml). The reaction mixture was stirred at room temperature (20–25°C) for 0.2–7 days (reaction completion was determined by disappearing of Sug-Cl according to TLC). The reaction mixture was filtered, the residue was washed with CHCl3/MeOH (1 : 1); the combined filtrate was evaporated, coevaporated with toluene, and dried in vacuo.

2.4. Deprotection and Separation

(see Table 1 for yields).

(a) Procedure for 4, 8, 15, and 17: the dry residue of treated reaction mixture (see above) was dissolved in 6 ml of dry MeOH and 0.3 ml 2 M MeONa/MeOH were added. The mixture was kept for 30 min at r.t., then evaporated followed by addition of 6 ml H2O. After 10–15 h (r.t.) the solution was evaporated, the residue was dissolved in ~1 ml water and the solution was applied on Dowex 50x4-400 (H+) ion-exchange resin column (  cm). The resin was washed sequentially with water (50 ml), 1 M aq. pyridine (50 ml) and 1 M aq. NH3 (50 ml). Target glycosides were completely eluted with 1 M pyridine. In the case of 10 and 17, nonreacted acceptor 16 was eluted with 1 M aq. NH3 in deprotected form (11). Pure α-glycoside 8 and corresponding β anomer were obtained by HPLC separation (ODS C18, water).

(b) Procedure for 10 was the same as (a), but included hydrogenolysis before MeONa/MeOH treatment: MeOH, 5% Pd/C, H2, 2 h. Pure α-glycoside 10 and corresponding β anomer were obtained by HPLC separation (ODS C18, water).

(c) Synthesis of 23 and 32 included procedure (a); products were eluted 1 M aq. pyridine. The obtained material was treated with 1 ml of 0.1 M NaOH for 2 h at r.t. to open lactone rings (partial lactonisation occurs during purification on Dowex H+) followed by neutralization with AcOH. Pure products as Na-salts were obtained after chromatography on silica gel (MeOH/MeCN/water, 3 : 3 : 1), purification on Sephadex LH-20 (MeCN/water, 1 : 1), cation exchange on Dowex 50x4-400 (Na+), and freeze drying.

(d) Procedure for 25 was the same as (c), with additional treatment with CF3COOH to remove Boc protection before the first Dowex chromatography. The mixture was dissolved in 0.3 ml CF3COOH, kept for 1 h at r.t., then coevaporated with toluene and dried in vacuo; the followed procedures were performed according to procedure (c).

(e) Synthesis of 24 included deprotection (treatment with MeONa/MeOH, NaOH/H2O, neutralization with AcOH), column chromatography on silica gel (2-propanol/MeCN/water 4 : 3 : 2), purification on Sephadex LH-20 (MeCN/water, 1 : 1), cation exchange on Dowex 50x4-400 (Na+) and freeze drying.

2.4.1. Characteristics of Synthesized Spacered Saccharides

1H NMR spectra were recorded at 30°C with a Varian 600 MHz instrument. Mass spectra (MALDI-TOF) were recorded on a Vision 2000 mass spectrometer. Optical rotations were measured with a JASCO DIP-360 polarimeter at 20°C. Thin-layer chromatography was performed using silica gel 60 F254 aluminum sheets (Merck, 1.05554) with detection by charring after 7% H3PO4 soaking, or ninhydrine.

9-Deoxy-9-NAc-Neu5Acα-O(CH2)3NH2 (15)
1H NMR spectrum (D2O, 303 K, 600 MHz)  ppm: 1.695 (1H, dd,  Hz, H3ax), 1.953 (2H, m, 2CH), 2.035 and 2.053 ( H, s, 2COCH3), 2.722 (1H, dd, 12,5, 4.7 Hz, H3eq), 3.138 (2H, m, 2CHN), 3.266 (1H, dd, 14.2, 7.7 Hz, H9), 3.504 (1H, dd, 1.4, 8.8 Hz, H7), 3.584 (1H, m, OCH), 3.629 (1H, dd, 14.2, 2.9 Hz, H9'), 3.704 (1H, ddd, 4.7, 11.8, 9.3 Hz, H4), 3.788 (1H, dd, 10.4, 1.4 Hz, H6), 3.839 (1H, dd, 10.4, 9.3 Hz, H5), 3.833 (1H, m, OCH), 3.903 (1H, ddd, 2.9, 7.7, 8.8 Hz, H8). MS, : 408 [M-]). +1.8 (c 1, H2O). 0.58 (MeOH/MeCN/water 3 : 3 : 2).

9-Deoxy-9-NAc-Neu5Acα2-6Galβ1-4GlcNAcβ-O(CH2)3NH2 (inner salt) (17)
NMR spectrum (D2O, 303 K, 600 MHz) δ ppm: 1.716 (1H, dd,  Hz, H3ax 9-deoxy-9-NAc-Neu5Ac), 1.975 (2H, m, 2CH), 2.037, 2.045, 2.092 ( H, s, 3COCH3), 2.677 (1H, dd, 4.5, 12.2 Hz, H3eq 9-deoxy-9-NAc-Neu5Ac), 3.114 (2H, m, 2NCH), 3.303 (1H, dd, 7.7, 14.0 Hz, H9 9-deoxy-9-NAc-Neu5Ac), 3.462 (1H, dd, 1.4, 9.0 Hz, H7 9-deoxy-9-NAc-Neu5Ac), 3.560 (2H, m, H2, H6 Galβ), 3.611 (1H, dd, 2.9, 14.0 Hz, H9′ 9-deoxy-9-NAc-Neu5Ac), 3.61–3.71 (4H, m, H4, H5 GlcNAcβ, 3.5, 9.8 Hz, H3 Galβ, H4 9-deoxy-9-NAc-Neu5Ac), 3.71–3.88 (7H, m, H2, H3 and H6 GlcNAcβ, H5 Galβ, H5, H6 9-deoxy-9-NAc-Neu5Ac, OCH), 3.945 (2H, m, H4 Galβ, H8 9-deoxy-9-NAc-Neu5Ac), 3.97–4.08, (3H, m, OCH, H6′ of GlcNAcβ, H6′ of Galβ), 4.464 (1H, d, 7.9 Hz, H1 Galβ), 4.560 (1H, m, H1 GlcNAcβ). MS, : 771 [M-]. −20(c 1, H2O). 0.41 (MeOH/MeCN/water 3 : 3 : 2).

Neu5Acα2-8Neu5Acα-O(CH2)3NH2 (Na+ salt) (23)
NMR spectrum (D2O, 303 K, 600 MHz) ppm: 1.646 (1H, dd, 12 Hz, H3ax Neu5Acα), 1.751 (1H, dd,  Hz, H3ax Neu5Acα2-8), 1.963 (2H, m, 2CH), 2.050 and 2.087 ( H, s, 2COCH3), 2.623 (1H, dd, 12.4, 4.4 Hz, H3eq Neu5Acα), 2.790 (1H, dd, 12.4, 4.5 Hz, H3eq Neu5Acα2-8), 3.148 (2H, m, 2CHN), 3.55–3.76 (7H, m, OCH, H4, H9 Neu5Acα, H4, H6, H7, H9 Neu5Acα2-8), 3.79–3.94 (7H, m, OCH, H5, H6, H7 Neu5Acα, H5, H8, H9′ Neu5Acα2-8), 4.113 (1H, dd. 3.9, 12.1 Hz, H9′ Neu5Acα), 4.198 (1H, m, H8 Neu5Acα). MS, : 723 [ ]. +3.2 (c 0.5, H2O). 0,44 (MeOH/MeCN/water 3 : 3 : 1).

Neu5Acα2-8Neu5Acα-OCH2(p-C6H4)NHCOCH2NH2 (Na+ salt) (25)
NMR spectrum (D2O, 303 K, 600 MHz) ppm: 1.651 (1H, dd, 12 Hz, H3ax Neu5Acα), 1.760 (1H, dd, 12 Hz, H3ax Neu5Acα2-8), 2.051 and 2.094 ( H, s, 2COCH3), 2.705 (1H, dd, 12.2, 4.3 Hz, H3eq Neu5Acα), 2.793 (1H, dd, 12.4, 4.6 Hz, H3eq Neu5Acα2-8), 3.58 – 3.75 (6H, m, H4, H9 Neu5Acα, H4, H6, H7, H9 Neu5Acα2-8), 3.81–3.95 (6H, m, H5, H6, H7 Neu5Acα, H5, H8, H9′ Neu5Acα2-8), 3.954 (2H, s, 2CHN), 4.188 (1H, dd, 12.1, 3.8, H9′ Neu5Acα), 4.247 (1H, m, H8 Neu5Acα), 4.504 and 4.504 ( H, d, 10.8 Hz, 2OCH-Ar), 7.478 (4H, m, C6H4). MS, : 763 [M-]). −17.0 (c 0.2, H2O). 0.60 (MeOH/MeCN/water 3 : 3 : 2).

Neu5Acα2-8Neu5Acβ-OCH2(p-C6H4)NHCOCH2NH2 (Na+ salt) (25β)
NMR spectrum (D2O, 303 K, 600 MHz) ppm: 1.620 (1H, dd, 12 Hz, H3ax Neu5Acα2-8), 1.690 (1H, dd, , H3ax Neu5Acβ), 2.035 and 2.105 ( H, s, 2COCH3), 2.391 (1H, dd, 13.2, 4.8 Hz, H3eq Neu5Acβ), 2.630 (1H, dd, 12.4, 4.7 Hz, H3eq Neu5Acα2-8), 3.52–3.64 (4H, m, H4, H6, H7, H9 Neu5Acα2-8), 3.685 (1H, dd, 12.1, , H9′ Neu5Acα2-8), 3.71–3.81 (4H, m, H5 Neu5Acα2-8, H6, H7, H9 Neu5Acβ), 3.87–3.94 (2H, m, H5 Neu5Acβ, H8 Neu5Acα2-8), 3.955 (2H, s, 2CHN), 4.083 (1H, ddd, H4 Neu5Acβ), 4.158 (2H, m, H8, H9′ Neu5Acβ), 4.355 and 4.542 ( H, 2d, 10.8 Hz, 2OCH-Ar), 7.527 (4H, m, C6H4). MS, : 763 [M-]). +10.0 (c 0.2, H2O). 0.40 (MeOH/MeCN/water 3 : 3 : 2).

Neu5Acα2-8Neu5Acα2-8Neu5Acα-O(CH2)3NH2 (Na+ salt) (32)
NMR spectrum (D2O, 303 K, 600 MHz) ppm: 1.652, 1.724 and 1.767 ( H, dd, , H3ax), 1.965 (2H, m, 2CH), 2.051, 2.089 and 2.097 ( H, s, 3COCH3), 2.621 (1H, dd, 4.7, 12.2 Hz, H3eq Neu5Acα), 2.701 (1H, dd, 12.0, 4.3 Hz, H3eq 2-8Neu5Acα2-8), 2.784 (1H, dd, 4.7, 12.2 Hz, H3eq Neu5Acα2-8), 3.150 (2H, m, 2CHN), 3.55–3.74 (10H, m, OCH, H4, H9; H6, H7 Neu5Acα2-8, H6 2-8Neu5Acα2-8), 3.79–3.97 (9H, m, H5; H6, H7 Neu5Acα, H7 2-8Neu5Acα2-8, H8, H9′ Neu5Acα2-8, OCH), 4.112 (3H, m, H8 Neu5Acα, H8, H9′ 2-8Neu5Acα2-8). MS, : 949 [M-]). +9.2 (c 1, H2O). 0.33 (MeOH/MeCN/water 3 : 3 : 1).

Neu5Acα2-8Neu5Acα2-8Neu5Acβ-O(CH2)3NH2 (Na+ salt) (32β)
NMR spectrum (D2O, 303 K, 600 MHz) δ ppm: 1.653, 1.674 and 1.739 ( H, dd, ), 2.015 (2H, m, 2CH), 2.050, 2.086 and 2.112 ( H, s, 3COCH3), 2.328 (1H, dd, 12.9, 4.9 Hz, H3eq Neu5Acβ), 2.730 (1H, dd, 12.4, 4.2 Hz, H3eq 2-8Neu5Acα2-8), 2.775 (1H, dd, 12.3, 4.5 Hz, H3eq Neu5Acα2-8), 3.130 (1H, m, NCH), 3.236 (1H, m, NCH), 3.386 (1H, m, OCH), 3.78–3.93 (6H, m, H5; H7 Neu5Acβ, H7 2-8Neu5Acα2-8, H9′ Neu5Acα2-8), 3.955 (1H, m, H8 Neu5Acα2-8), 4.009 (1H, ddd, 4.8, 11.0 Hz, H4 Neu5Acβ), 4.064 (1H, dd, 12.3, 2.6 Hz, H9′ Neu5Acβ), 4.096 (1H, m, H8 Neu5Acβ), 4.159 (2H, m, H8, H9′ 2-8Neu5Acα2-8). MS, : 949 [M-]). +9.0 (c 0.3, H2O). 0.22 (MeOH/MeCN/water 3 : 3 : 1).


This work was supported with FP6 (project “Avian influenza: impact of virus-host interactions on pathogenesis and ecology”, acronym: FLUPATH, contract No. 044220) and Grant of Russian Foundation for Basic Research 07-04-00630.


  1. R. Schauer, “Achievements and challenges of sialic acid research,” Glycoconjugate Journal, vol. 17, no. 7-9, pp. 485–499, 2000. View at: Publisher Site | Google Scholar
  2. A. Varki, “Sialic acids in human health and disease,” Trends in Molecular Medicine, vol. 14, no. 8, pp. 351–360, 2008. View at: Publisher Site | Google Scholar
  3. G.-J. Boons and A. V. Demchenko, “Recent advances in O-sialylation,” Chemical Reviews, vol. 100, no. 12, pp. 4539–4565, 2000. View at: Publisher Site | Google Scholar
  4. R. L. Halcomb and M. D. Chappell, “Recent developments in technology for glycosylation with sialic acid,” Journal of Carbohydrate Chemistry, vol. 21, no. 7–9, pp. 723–768, 2002. View at: Publisher Site | Google Scholar
  5. Y. Liu, X. Ruan, X. Li, and Y. Li, “Efficient synthesis of a sialic acid α(2→3)galactose building block and its application to the synthesis of ganglioside GM3,” Journal of Organic Chemistry, vol. 73, no. 11, pp. 4287–4290, 2008. View at: Publisher Site | Google Scholar
  6. G. Pazynina, A. Tuzikov, A. Chinarev, P. Obukhova, and N. Bovin, “Simple stereoselective synthesis of α2-6 sialooligosaccharides,” Tetrahedron Letters, vol. 43, no. 45, pp. 8011–8013, 2002. View at: Publisher Site | Google Scholar
  7. T. Yamaji, T. Teranishi, M. S. Alphey, P. R. Crocker, and Y. Hashimoto, “A small region of the natural killer cell receptor, Siglec-7, is responsible for its preferred binding to α2,8-disialyl and branched α2,6-sialyl residues. A comparison with Siglec-9,” Journal of Biological Chemistry, vol. 277, no. 8, pp. 6324–6332, 2002. View at: Publisher Site | Google Scholar
  8. L. A. Simeoni, A. B. Tuzikov, N. E. Byramova, and N. V. Bovin, “Neu5Ac and Neu5Gc ethylthioglycoside derivatives as glycosyl donors. Synthesis of aminopropyl glycosides of N-acetyl and N-glycolylneuraminic acid,” Russian Journal of Bioorganic Chemistry, vol. 23, pp. 139–146, 1997. View at: Google Scholar
  9. A. Hasegawa, H. Ishida, and M. Kiso, “Synthetic studies on sialoglycoconjugates 46: a facile total synthesis of ganglioside GD3,” Journal of Carbohydrate Chemistry, vol. 12, pp. 371–376, 1993. View at: Google Scholar
  10. H. Ando, H. Ishida, M. Kiso, and A. Hasegawa, “A synthetic approach to the c-series gangliosides containing sialyl-α(2 → 8) sialyl-α(2 → and) sialic acid: synthesis of ganglioside GT4, α(2 → 6) GT4 and GT3,” Carbohydrate Research, vol. 300, no. 3, pp. 207–217, 1997. View at: Publisher Site | Google Scholar
  11. A. B. Tuzikov, A. S. Gambaryan, L. R. Juneja, and N. V. Bovin, “Conversion of complex sialooligosaccharides into polymeric conjugates and their anti-influenza virus inhibitory potency,” Journal of Carbohydrate Chemistry, vol. 19, no. 9, pp. 1191–1200, 2000. View at: Google Scholar
  12. S. Z. Abbas, S. Sugiyama, J. Diakur, R. A. Pon, and R. Roy, “Synthesis of α- and β-methyl Neu5Acα-(2→8)Neu5Ac disaccharides,” Journal of Carbohydrate Chemistry, vol. 9, no. 6, pp. 891–901, 1990. View at: Google Scholar
  13. A. A. Sherman, O. N. Yudina, A. S. Shashkov, V. M. Menshov, and N. E. Nifant'ev, “Synthesis of Neu5Ac- and Neu5Gc-α-(26)-lactosamine 3-aminopropyl glycosides,” Carbohydrate Research, vol. 330, no. 4, pp. 445–458, 2001. View at: Publisher Site | Google Scholar
  14. H. J. Gross, A. Bünsch, J. C. Paulson, and R. Brossmer, “Activation and transfer of novel synthetic 9-substituted sialic acids,” European Journal of Biochemistry, vol. 168, no. 3, pp. 595–602, 1987. View at: Google Scholar
  15. F. Michon, J.-R. Brisson, and H. J. Jennings, “Conformational differences between linear α(28)-linked homosialooligosaccharides and the epitope of the group B meningococcal polysaccharide,” Biochemistry, vol. 26, no. 25, pp. 8399–8405, 1987. View at: Google Scholar
  16. E. M. Rapoport, G. V. Pazynina, M. A. Sablina, P. R. Crocker, and N. V. Bovin, “Probing sialic acid binding Ig-like lectins (siglecs) with sulfated oligosaccharides,” Biochemistry, vol. 71, no. 5, pp. 496–504, 2006. View at: Publisher Site | Google Scholar
  17. O. A. Kost, N. V. Bovin, E. E. Chemodanova, V. V. Nasonov, and T. A. Orth, “New feature of angiotensin-converting enzyme: carbohydrate-recognizing domain,” Journal of Molecular Recognition, vol. 13, no. 6, pp. 360–369, 2000. View at: Google Scholar
  18. E. M. Rapoport, YU. B. Sapot'ko, G. V. Pazynina, V. K. Bojenko, and N. V. Bovin, “Sialoside-binding macrophage lectins in phagocytosis of apoptotic bodies,” Biochemistry, vol. 70, no. 3, pp. 330–338, 2005. View at: Publisher Site | Google Scholar
  19. R. Brossmer and H. J. Gross, “Sialic acid analogs and application for preparation of neoglycoconjugates,” Methods in Enzymology, vol. 247, pp. 153–176, 1994. View at: Publisher Site | Google Scholar

Copyright © 2010 Galina Pazynina et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

More related articles

 PDF Download Citation Citation
 Download other formatsMore
 Order printed copiesOrder

Related articles

Article of the Year Award: Outstanding research contributions of 2020, as selected by our Chief Editors. Read the winning articles.