An efficient synthesis of D-galactose-based multivalent neoglycoconjugates

ARTICLE

An efficient synthesis of D-galactose-based multivalent neoglycoconjugates

Saulo F. de Andrade^I; Rute C. Figueiredo^III; José D. de Souza Filho^I; Ricardo J. AlvesII, ^*

^IDepartamento de Química, Instituto de Ciências Exatas

^IIFaculdade de Farmácia, Universidade Federal de Minas Gerais, Av. Antônio Carlos, 6627, 31270-901 Belo Horizonte-MG, Brazil

^IIIUniversidade Federal de Ouro Preto, Campus Morro do Cruzeiro, s/n, Bairro Bauxita, 35400-000 Ouro Preto-MG, Brazil

ABSTRACT

In this work, the synthesis of dimeric, trimeric and tetrameric D-galactose-based neoglycoconjugates is reported. The monosaccharide ligand was prepared in 5 straightforward steps from D-galactose using the Doebner modification of the Knoevenagel reaction for chain elongation. The ligand was coupled to 1,4-butanediamine, tris-(2-ethylamino)amine, pentaerythrityltetramine and PAMAM dendrimers (1,4-butanodiamine core G0 and 1,12-dodecanediamine core G0). The unprotected glycodendrimers were purified by size-exclusion chromatography (SEC). This was the only step in which a chromatographic method was employed throughout the synthetic route. This is a new and efficient strategy for the preparation of neoglycoconjugates.

Keywords: glycodendrimers, D-galactose, synthesis, pentaerythrityltetramine, PAMAM

RESUMO

Este trabalho descreve a síntese de neoglicoconjugados diméricos, triméricos e tetraméricos derivados de D-galactose. O ligante monossacarídico foi preparado em cinco etapas utilizando-se a modificação de Doebner da reação de Knoevenagel na etapa de extensão da cadeia lateral. O ligante foi acoplado a 1,4-butanodiamina, tris-(2-etilamino)amina, pentaeritritiltetramina e dendrímeros PAMAM (núcleo 1,4-butanodiamina G0 e núcleo 1,12-dodecanodiamina G0). Os glicodendrímeros desprotegidos foram purificados por cromatografia por exclusão de tamanho (SEC). Esta foi a única etapa em que um método cromatográfico foi empregado em toda rota sintética. Essa é uma estratégia nova e eficiente para a preparação de neoglicoconjugados.

Introduction

Glycodendrimers are neoglycoconjugates that can be considered as bioisosters of glycoproteins,¹ as they can mimic the multivalent interactions of lectin-carbohydrate in several adhesion processes. The ability of glycodendrimers to present multivalent interactions with lectins as compared to a monovalent ligand is referred to as "cluster effect".^2,3 Increasing multivalency, however, does not always result in higher affinity. Several low molecular weight and low valence glycodendrimers (dimers, trimers and tetramers) are very potent and, sometimes, even more potent than higher valency analogs in their interaction with lectins.^4-7

This is very important from the medicinal chemistry point of view, since low molecular dendrimers are expected to have favorable physicochemical properties comparing to the high molecular weight, multivalent ones.

Efforts for blocking microbial lectins have driven carbohydrate chemists towards the production of original molecules specifically designed for their anti-adhesion potency. For example, L-fucoside and D-galactose-coated glycodendrimers were effective in blocking P. aeruginosa lectin PA-IL and PA-IIL,⁸ while D-mannose-coated ones were able to inhibit the binding of E. coli to epithelial cells.⁹ Recently, anti-inflammatory activity was reported for D-glucose-based glycodendrimers.¹⁰

Due to the importance of neoglycoconjugates for these biological studies, there is a constant need for straightforward methods for the synthesis and purification of these compounds. Herein, we present an efficient synthesis of the glycodendrimers 1-5 (Figure 1).

Results and Discussion

Amide bonds are one of the most useful connections to attach a carbohydrate moiety to a core or dendrimer.¹¹ Hence, the preparation of a carboxylic acid glycoside is essential to couple with polyamine cores or dendrimers. Glycodendrimers 1-5 were obtained from the galactopyranoside 9 and the commercially available cores 1,4-butanediamine and tris-(2-ethylamino)amine; commercially available dendrimers PAMAM (1,4-butanodiamine core G0 and 1,12-dodecanediamine core G0) and synthetic pentaerythrityltetramine (Scheme 1). The monosaccharide ligand 9 was easily prepared on multigram scale in 4 straightforward steps from peracetylated D-galactose. Compound 10 was coupled to the cores or dendrimers followed by the methanolysis.¹² This afforded the glycodendrimers 1-5.

Pentaerythrityltetramine was prepared in the form of the corresponding tetrahydrochloride from pentaerythritol by a modification of previous report in three steps (Scheme 2).¹³

Preparation of glycoside 9

The known compound 8 was prepared in 3 steps by a modification of previous report.¹⁴ Initially, the treatment of peracetylated galactose with HBr provided the galactopyranosyl bromide 6.¹⁵ Glycosylation of the inexpensive and commercially available vanillin with 6, under basic conditions, furnished the glycoside 7 (68% overall yield after recrystallization).¹⁶ Glycoside 7 was reacted with malonic acid under Doebner conditions to give the known compound 8 in 96% yield.^17,18

Finally, hydrogenation of 8 using H₂/Pd-C afforded the novel glycoside 9 in a pure form (90% yield).¹⁸ In the ¹H NMR (nuclear magnetic resonance) spectrum of 9 (Figure 2), the signals of the aromatic protons were detected at δ 7.05-6.72. Galactopyranoside ring proton signals were observed at d 5.49-3.97. The protons of the methoxy group were observed at δ 3.81, methylene units at d 2.91-2.66 and methyls of the acetate groups at δ 2.18-2.01. Thus, compound 9 could be easily prepared in pure form on a gram scale without using any column chromatography.

Preparation of glycodendrimers 1-5

The carboxy group of 9 was activated with N-hydroxysuccinimide (NHS) using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC) to give 10.¹⁹ The reaction of 10 in small excess (1.2 equivalent per mol of amino group) with cores or dendrimers followed by O-deacetylation under Zemplén's conditions gave glycodendrimers 1-5 in 75-94% yield, as described in the Experimental section.²⁰ The excess of activated ester 10 was removed by size exclusion chromatography (SEC) performed using MeOH/H₂O (1:1) as mobile phase. The Sephadex column could be efficiently recycled by washing with 100 mL of the mobile phase.

Glycodendrimers characterization

Here, the analysis of the most complex structure glycodendrimer 5 is exemplified. In the ¹H NMR of 5 (Figure 3), the assignments of the NHC=O (δ 7.92-7.86), H-1 (δ 4.78), H-9 (δ 6.81) OCH₃ (δ 3.73), H-4 (δ 3.69), H-3 (δ 3.38), Ha and Hb (δ 3.07), Hf (δ 1.33) and Hg-j (δ 1.07) were made based on chemical shift, coupling constant, multiplicity and integral values.

Comparing the integral of galactopyranoside proton signals (e.g. H-1) with dendrimer proton signals (e.g. Ha), it was possible to establish the proportion of the sugar units related to the dendrimer nucleus (1:2 ratio in this case). Besides, the HMBC contour map allowed establishing unequivocally the correlation between the carbonyl carbon of the carbohydrate residue with Ha protons.

The assignments for H-2 and He were based on COSY contour map. Correlations between H-1 (δ 4.78) /H-2 (d 3.56) and Hf (δ 1.33) /He (2.36-2.32) were observed. Besides, most carbons were assigned using HSQC. Cross-peaks between H-1 (d 4.78)/ C-1 (δ 100.9), H-2 (δ 3.56)/C-2 (δ 70.3), H-3 (δ 3.38)/C-3 (δ 73.5), H-4 (d 3.69)/C-4 (δ 68.0), H-9 (δ 6.81)/C-9 (δ 122.8), Ha (d 3.07)/Ca (38.3), Hb (d 3.07)/Cb (38.3), He (δ 2.36-2.32)/Ce (δ 52.8), Hf (δ 1.33)/Cf (δ 26.6), Hg-Hj (δ 1.22)/Cg-Cj (d 27.0) and OCH₃ (d 3.73)/OCH₃ (d 55.6) were detected. HMBC (heteronuclear multiple bond correlation) map was used to assign non-hydrogenated carbons C-7, C-8 and C-10. Some important ³J correlations were observed between H-1 (δ 4.78)/C-7 (δ 144.8), OCH₃ (δ 3.73)/C-8 (d 148.8) and H-14 (δ 2.36-2.32)/C-10 (135.0)..

The analysis by electrospray ionization mass spectroscopy (ESI-MS) was also conducted. Interestingly, the ESI-MS spectra of compounds 3-5 give peaks with fractional values (m/z; z = 2). This is assigned to dicationic species [M + 2Na⁺] for compounds 3-4 and [M + 2H⁺] for 5. This is a common fact in ESI-MS.²¹

2D spectra (COSY, HSQC and HMBC) and mass spectrum are available in the Supplementary Information (SI) section.

Conclusions

An efficient access to glycodendrimers was demonstrated in this work. The synthesis was performed in few steps using cheap reactants in 44-55% overall yields. This synthetic strategy can be extended to other sugars and aromatic aldehydes as a general method of obtaining low molecular weight glycodendrimers.

Experimental

Material and methods

Melting points were determined on Microquímica MQAPF 301 apparatus. IR spectra were recorded on a Spectrum One, Perkin-Elmer ATR system. The ¹H and ¹³C NMR spectra were obtained on a Bruker Avance DPX-200 or DRX-400. The proton and carbon chemical shifts (d) are given with respect to TMS (tetramethylsilane). The assignments were based on COSY, DEPT-135, HSQC and HMBC experiments. Optical rotations were measured on an ADP220 Bellinghan + Stanley Ltd polarimeter. Mass spectra were recorded with a Waters Quattro LC triple quadrupolar mass spectrometer. Thin layer chromatography was performed using silica gel 60G Merck and 15% ethanolic H₂SO₄ (v/v) was employed for the revelation of the spots. Size-exclusion chromatography (SEC) was performed using Sephadex LH-20 (25-100 µm). The reagents were obtained from commercial suppliers and were used without further purification.

4-Formyl-2-methoxyphenyl 2,3,4,6-tetra-O-acetyl-β-D-galactopyranoside 7

A 48% of an HBr aqueous solution (m/m; 24.7 mL, 0.218 mol) was added to an ice-cold magnetically stirred acetic anhydride (100 mL, 1.06 mol) by means of an addition funnel. The resulting solution was added to an ice-cold magnetically stirred solution of 1,2,3,4,6-penta-O-acetyl-α,β-galactopyranoside (10 g, 25.6 mmol) in dichloromethane (75 mL) by means of an addition funnel. The mixture was warmed to room temperature. After 2 h, TLC analysis (9:1 CH₂Cl₂/EtOAc) showed that the reaction was complete. The mixture was extracted with dichloromethane (3 × 50 mL). The combined organic layers were washed with a saturated aqueous solution of sodium bicarbonate to remove the acid followed by the saturated aqueous brine solution. The organic phase was dried with sodium sulfate, filtered and concentrated to give 2,3,4,6-tetra-O-acetyl-α-D-galactopyranosyl bromide 6 as a colorless oil (10.61 g, 100% yield) which was used without further purification.

To a solution of LiOH.H₂O (3.06 g, 73 mmol) in water (23 mL), it was added 4-hydroxy-3-methoxybenzaldehyde (12.16 g, 80 mmol) and the mixture was stirred for 10 min. Then, a solution of 6 (10.52 g) in acetone (30 mL) was added with stirring. After 2 h, TLC analysis (9:1 CH₂Cl₂/EtOAc) showed the completion of the reaction. Later, acetone was removed under reduced pressure and water (50 mL) was added to the resulting mixture. The aqueous phase was extracted with dichloromethane (2 × 50 mL). The combined organic layers were washed with 1 mol L^-1 aqueous NaOH (3 × 100 mL) and with water to pH 7. The organic phase was dried with anhydrous sodium sulfate, filtered and concentrated. The crude product was dissolved in acetone (5 mL), water (200 mL) was added and the solid formed was collected by filtration to give 7 as a white solid (8.52 g, 68% yield, 2 steps): mp 123-124 ºC; [α]_D²¹ -24.0 (c 1.0, MeOH); IR ν/cm^-1 2867 and 1693 (aldehyde) 1752, 1740 and 1224 (ester) 1588 and 1515 (Aryl); ¹H NMR (CDCl₃, 200 MHz) d 9.89 (s, 1H, CHO), 7.44-7.41 (m, 2H, H-9 and H-11), 7.24 (d, 1H, J_11,12 8.4 Hz, H-12), 5.56 (dd, 1H, J_1,2 8.0 Hz and J_2,3 10.2 Hz, H-2), 5.47 (d, 1H, J_3,4 3.2 Hz, H-4), 5.13 (dd, 1H, J_2,3 10.2 Hz and J_3,4 3.2 Hz, H-3), 5.07 (d, 1H, J_1,2 8.0 Hz, H-1), 4.30-4.05 (m, 3H, H-5, H-6 and H-6'), 3.90 (s, 3H, OCH₃), 2.18, 2.08, 2.06, 2.02 (s, 12H, COCH₃); ¹³C NMR (CDCl₃; 50 MHz) δ 190.8 (CHO); 170.2; 170.1; 169.3 (COCH₃); 151.2; 150.8 (C-7 and C-8); 132.6 (C-10); 125.3 (C-11); 117.7 (C-9); 110.6 (C-12); 100.2 (C-1); 71.1 (C-3); 70.4 (C-5); 68.3 (C-2); 66.7 (C-4); 61.2 (C-6); 56.0 (OCH₃); 20.5 (COCH₃).

4-E-[(2-Carboxy)ethenyl]-2-methoxyphenyl 2,3,4,6-tetra-O-acetyl- β -D-galactopyranoside 8

A solution of 7 (2.0 g, 4.15 mmol), malonic acid (2.6 g, 25 mmol) in a mixture of pyridine (6.5 mL) and pyperidine (0.5 mL) was stirred at 95 ºC. After 3 h, TLC analysis (1:1 hexane/EtOAc) showed that the reaction was complete. The mixture was cooled in an ice-water bath and acidified to pH 1-2 by the slow addition of 6 mol L^-1 aqueous HCl. The resulting suspension was extracted with dichloromethane (2 × 50 mL). The combined organic layers were washed with water to pH 7, dried with sodium sulfate, filtered and concentrated to afford a amorphous white solid (2.17 g, 96% yield): mp 77-80 ºC; [α]_D²¹ -10 (c 1.0, CHCl₃); IR ν/cm^-1 3000 and 1686 (carboxylic acid), 1744 (ester), 1631 (olefin); ¹H NMR (CDCl₃, 200 MHz) d 7.71 (d, 1H, J_13,14 15.8 Hz, H-13), 7.12-7.09 (m, 3H, H-9, H-11 and H-12), 6.35 (d, 1H, J_13,14 15.8 Hz, H-14), 5.53 (dd, 1H, J_1,2 8.0 Hz and J_2,3 10.4 Hz, H-2), 5.45 (d, 1H, J_3,4 3.1 Hz, H-4), 5.12 (dd, 1H, J_2,3 10.4 Hz and J_3,4 3.1 Hz, H-3), 4.98 (d, 1H, J_1,2 8.0 Hz, H-1), 4.35-4.00 (m, 3H, H-5, H-6 and H-6'), 3.86 (s, 3H, OCH₃), 2.18, 2.09, 2.05, 2.02 (4s, 12H, COCH₃); ¹³C NMR (CDCl₃; 50 MHz) δ 171.9; 170.6; 170.5; 169.7 (COCH₃ and COOH); 150.8 (C-8); 148.5 (C-7); 146.4 (C-13); 130.6 (C-10); 122.3 (C-12); 119.2 (C-11); 116.7 (C-14); 111.7 (C-9); 100.9 (C-1); 71.2; 70.8 (C-3 and C-5); 68.7 (C-2); 67.0 (C-4); 61.5 (C-6); 56.2 (OCH₃); 20.8 (COCH₃).

4-(2-Carboxyethyl)-2-methoxyphenyl 2,3,4,6-tetra-O-acetyl-β-D-galactopyranoside 9

To solution of 8 (0.5 g, 0.95 mmol) in THF (tetrahydrofuran, 5.0 mL), it was added Pd/C (0.1 g). The mixture was stirred under hydrogen atmosphere (1 atm) for 24 h. The catalyst was removed by filtration and the solvent evapored under reduced pressure to afford a white solid (0.45 g, 90% yield): mp 77-78 ºC; [α]_D²¹ -8 (c 1.0, CHCl₃); IR ν/cm^-1 3000 and 1743 (carboxylic acid), 1743 (ester); ¹H NMR (CDCl₃, 200 MHz) d 7.05 (d, 1H, J_11,12 8.1 Hz, H-12), 6.76 (s, 1H, H-9), 6.72 (d, 1H, J_11,12 8.1 Hz, H-11), 5.49 (dd, 1H, J_1,2 7.9 Hz and J_2,3 10.5 Hz, H-2), 5.44 (d, 1H, J_3,4 3.4 Hz, H-4), 5.1 (dd, 1H, J_2,3 10.5 Hz and J_3,4 3.4 Hz, H-3), 4.88 (d, 1H, J_1,2 7.9 Hz, H-1), 4.29-4.1 (m, 2H, H-6 and H-6'), 3.97 (t, 1H, J_5,6 6.6 Hz, H-5), 3.81 (s, 3H, OCH₃), 2.91 (t, 2H, J_13,14 7.4 Hz, H-13), 2.66 (t, 2H, J_13,14 7.4 Hz, H-14), 2.18, 2.09, 2.04, 2.01 (4s, 12H, COCH₃); ¹³C NMR (CDCl₃; 50 MHz) δ 178.0 (COOH); 170.4; 170.3; 170.2; 169.5 (COCH₃); 150.4 (C-8); 144.6 (C-7); 136.8 (C-10); 120.2; 120.0 (C-11 and C-12); 112.8 (C-9); 101.3 (C-1); 70.7 (C-3 and C-5); 68.7 (C-2); 66.9 (C-4); 61.2 (C-6); 55.9 (OCH₃); 35.5 (C-14); 30.2 (C-13); 20.5 (COCH₃).

General procedure for the synthesis of D-galactose-based dendrimers

A suspension of 9 (1.0 g, 1.90 mmol), EDC (0.6 g, 3.04 mmol), NHS (0.350 g, 3.04 mmol) in dichloromethane (20 mL) was stirred at room temperature. After 3 h, TLC analysis (3:7 hexane/EtOAc) showed that the reaction was complete. To the solution, it was added dichloromethane (30 mL). The organic phase was washed with brine (2 × 50 mL), dried with sodium sulfate and concentrated to give 4-[[[2-succinimid-1-yl)oxy]carbonyl]ethyl]-2-methoxyphenyl 2,3,4,6-tetra-O-acetyl-β-D-galactopyranoside 10 as a colorless foam (1.17 g) which was used without further purification: IR ν/cm^-1 1734 and 1210 (ester) 1594 (Aryl); ¹H NMR (CDCl₃, 200 MHz) δ 7.07 (d, 1H, J_11,12 8.1 Hz, H-12), 6.77 (s, 1H, H-9), 6.73 (d, 1H, J_11,12 8.1 Hz, H-11), 5.49 (dd, 1H, J_1,2 8.0 Hz and J_2,3 10.6 Hz, H-2), 5.43 (d, 1H, J_3,4 3.5 Hz, H-4), 5.09 (dd, 1H, J_2,3 10.6 Hz and J_3,4 3.5 Hz, H-3), 4.88 (d, 1H, J_1,2 8.0 Hz, H-1), 4.28-4.10 (m, 2H, H-6 and H-6'), 3.97 (t, 1H, H-5, J_5,6 6.5 Hz), 3.82 (s, 3H, OCH₃), 2.98 (t, 2H, J_13,14 6.2 Hz, H-13); 2.91 (t, 2H, J_13,14 6.2 Hz, H-14), 2.84 (s, 4H, H-17), 2.18, 2.09, 2.04, 2.01 (s, 12H, COCH₃); DEPT-135 ¹³C NMR (CDCl₃; 50 MHz): δ 120.2; 120.1 (C-11 and C-12); 112.7 (C-9); 101.3 (C-1); 70.7; 70.6 (C-3 and C-5); 68.6 (C-2); 66.8 (C-4); 61.2 (C-6); 55.9 (OCH₃); 32.6 (C-14); 30.1 (C-13); 25.5 (C-17); 20.5 (COCH₃).

To the pure core or dendrimer methanol solution (in this case the methanol was co-destilled with dichloromethane, 3 × 5 mL) was added dichloromethane (5 mL). To the resulting solution, it was added a solution of 10 (0.1 g, 0.16 mmol, 1.2 molar equivalent per terminal amino group) in dichloromethane (5 mL) with stirring at room temperature. In some cases, the formation of a suspension was observed after about 1 h. In these cases, methanol was dropwise until a solution was obtained. After 3 h, the solvent was removed and the crude was dissolved in methanol. The mixture was cooled in an ice-water bath and a solution of sodium methoxide in methanol was added. The mixture was stirred for 1 h, then acidified to pH 7 by the addition of 1 mol L^-1 aqueous HCl and concentrated under reduced pressure. The residue was purified by SEC (1:1 MeOH/H₂O).

Glycodendrimer 1

1,4-Butanediamine (5.8 mg, 0.067 mmol) was used as core. Colorless semi-solid (48 mg, 94% yield): IR ν/cm^-1 3675-2992 (amide and alcohol), 1637 and 1554 (amide), 1512 (Aryl); ¹H NMR (DMSO-d₆, 400 MHz) d 7.76 (t, 2H, J_a,NH 5.4 Hz, NH's), 6.96 (d, 2H, J_11,12 8.5 Hz, H-12), 6.81 (d, 2H, J_9,11 1.6 Hz, H-9), 6.67 (dd, 2H, J_9,11 1.6 Hz and J_11,12 8.5 Hz, H-11), 5.01 (bs, 2H, OH-2), 4.84 (bs, 2H, OH-6), 4.79 (d, 2H, J_1,2 8.0 Hz, H-1), 4.61 (bs, 2H, OH-3), 4.50 (bs, 2H, OH-4), 3.73 (s, 6H, OCH₃), 3.69 (bs, 2H, H-4), 3.53 (m, 2H, H-2), 3.50-3.44 (m, 6H, H-5, H-6 and H-6'), 3.38 (dd, 2H, J_2,3 9.6 Hz and J_3,4 2.8 Hz, H-3), 3.01 (bq, 4H, J_a,NH = J_a,b 5.4, Ha), 2.74 (t, 4H, J_13,14 7.8 Hz, H-13), 2.33 (t, 4H, J_13,14 7.8 Hz, H-14), 1.33 (bs, 4H, H-b); ¹³C NMR (DMSO-d₆; 100 MHz) d 171.2 (NHC=O); 148.8 (C-8); 144.8 (C-7); 135.1 (C-10); 120.0 (C-11); 115.5 (C-12); 112.8 (C-9); 100.9 (C-1); 75.4 (C-5); 73.5 (C-3); 70.3 (C-2); 68.1 (C-4); 60.3 (C-6); 55.6 (OCH₃); 38.1 (Ca); 37.3 (C-14); 30.8 (C-13); 26.6 (Cb); MS-ESI m/z calcd. for C₃₆H₅₂N₂O₁₆ 768.8, found 792.0 (M + Na⁺).

Glycodendrimer 2

Tris(2-aminoethyl)amine (6.5 mg, 0.044 mmol) was used as core. Amorphous white solid (39 mg, 75% yield): IR ν/cm^-1 3675-3002 (amide and alcohol), 1636 and 1558 (amide), 1510 (Aryl); ¹H NMR (DMSO-d₆, 400 MHz) d 7.76 (t, 3H, J_a,NH 5.7 Hz, NH's), 6.98 (d, 3H, J_11,12 8.4 Hz, H-12), 6.83 (d, 3H, J_9,11 1.6 Hz, H-9), 6.69 (dd, 3H, J_9,11 1.6 Hz and J_11,12 8.4 Hz, H-11), 5.2-4.3 (m, OH's), 4.81 (d, 3H, J_1,2 7.6 Hz, H-1), 3.74 (s, 9H, OCH₃), 3.72 (d, 3H, J_3,4 3.0 Hz, H-4), 3.59 (m, 3H, H-2); 3.55-3.45 (m, 9H, H-5, H-6 and H-6'); 3.41 (dd, 3H, J_2,3 9.2 Hz and J_3,4 3.0 Hz, H-3), 3.10 (bq, 6H, J_a,NH = J_a,b 5.7 Hz, Ha), 2.77 (t, 6H, J_13,14 7.9 Hz, H-13), 2.48 (t, 6H, J_a,b 5.7 Hz, Hb), 2.39 (t, 6H, J_13,14 7.9 Hz, H-14); ¹³C NMR (DMSO-d₆; 100 MHz) δ 171.4 (NHC=O); 148.8 (C-8); 144.8 (C-7); 135.0 (C-10); 119.9 (C-11); 115.4 (C-12); 112.8 (C-9); 100.8 (C-1); 75.3 (C-5); 73.5 (C-3); 70.3 (C-2); 68.0 (C-4); 60.3 (C-6); 55.6 (OCH₃); 53.5 (Cb); 37.2 (C-14); 36.9 (Ca); 30.7 (C-13); MS-ESI m/z calcd. for C₅₄H₇₈N₄O₂₄ 1166.2, found 1167.3 (M + H⁺).

Glycodendrimer 3

Pentaerythrityltetramine was obtained from the corresponding tetrahydrochloride (9.2 mg, 0.033 mmol) and sodium bicarbonate (29 mg, 0.29 mmol). Dichloromethane was replaced by acetone (5 mL) and water (sufficient amount to dissolve the reactants). Amorphous white solid (38 mg, 78% yield): IR ν/cm^-1 3691-3022 (amide and alcohol), 1631 and 1541 (amide), 1511 (Aryl); ¹H NMR (DMSO-d₆, 400 MHz) d 7.68-7.59 (m, 4H, NH's), 6.97 (d, 4H, J_11,12 8.4 Hz, H-12), 6.85 (s, 4H, H-9), 6.70 (d, 4H, J_11,12 8.4 Hz, H-11), 4.98 (d, 4H, J_OH-2,2 4.8 Hz, OH-2), 4.80-4.78 (m, 8H, J_1,2 7.6 Hz, OH-6 and H-1), 4.59 (bs, 4H, OH-3), 4.45 (d, 4H, J_OH-4,4 4.0 Hz, OH-4), 3.73 (s, 12H, OCH₃), 3.67 (bs, 4H, H-4), 3.59-3.39 (m, 20H, H-2, H-3, H-5, H-6 and H-6'), 2.80 (t, 8H, J_13,14 7.4 Hz, H-13), 2.72-2.66 (m, 8H, Ha), 2.46 (t, 8H, J_13,14 7.4 Hz, H-14); ¹³C NMR (DMSO-d₆; 100 MHz) δ 172.6 (NHC=O); 148.9 (C-8); 144.9 (C-7); 134.6 (C-10); 120.0 (C-11); 115.5 (C-12); 112.8 (C-9); 100.9 (C-1); 75.3 (C-5); 73.5 (C-3); 70.3 (C-2); 68.1 (C-4); 60.3 (C-6); 55.6 (OCH₃); 39.0 (Ca); 37.5 (C-14); 30.6 (C-13); MS-ESI m/z calcd. for C₆₉H₉₆N₄O₃₂ 1493.5, found 769.7 (M + 2Na⁺).

Glycodendrimer 4

A 20% (w/w) methanol solution of PAMAM dendrimer, 1,4-butanodiamine core, G0 (90 mg of solution, 0.033 mol) was used. Crystalline solid (50 mg, 80% yield): IR ν/cm^-1 3691-2997 (amide and alcohol), 1635 and 1550 (amide), 1512 (Aryl); ¹H NMR (DMSO-d₆, 400 MHz) d 7.97-7.92 (m, NH's), 6.98 (d, 4H, J_11,12 8.4 Hz, H-12), 6.83 (d, 4H, J_9,11 1.6 Hz, H-9), 6.69 (dd, 4H, J_9,11 1.6 Hz and J_11,12 8.4 Hz, H-11), 5.10-4.45 (m, OH's), 4.81 (d, 4H, J_1,2 8.0 Hz, H-1), 3.75 (s, 12H, OCH₃), 3.71 (d, 4H, J_3,4 3.2 Hz, H-4), 3.57 (m, 4H, H-2), 3.54-3.44 (m, 12H, H-5, H-6 and H-6'), 3.41 (dd, 4H, J_2,3 9.6 Hz and J_3,4 3.2 Hz, H-3), 3.09 (bs, 16H, Ha and Hb), 2.76 (t, 8H, J_13,14 7.8 Hz, H-13), 2.63 (t, 8H, J_c,d 6.8 Hz, Hd), 2.38-2.34 (m, 12H, He and H-14), 2.20 (t, 8H, J_c,d 6.8 Hz, Hc), 1.34 (bs, 4H, Hf); ¹³C NMR (DMSO-d₆; 100 MHz) d 171.6 (NHC=O's); 148.8 (C-8); 144.8 (C-7); 135.0 (C-10); 120.0 (C-11); 115.4 (C-12); 112.8 (C-9); 100.9 (C-1); 75.3 (C-5); 73.5 (C-3); 70.3 (C-2); 68.0 (C-4); 60.3 (C-6); 55.6 (OCH₃); 52.7 (Ce); 49.3 (Cd); 38.3 (Ca and Cb); 37.3 (C-14); 33.2 (Cc); 30.7 (C-13); 24.3 (Cf); MS-ESI m/z calcd. for C₈₈H₁₃₂N₁₀O₃₆ 1906.0, found 976.2 (M + 2Na⁺).

Glycodendrimer 5

A 20% (w/w) methanol solution of PAMAM dendrimer, 1,12-dodecanediamine core, G0 (108 mg of solution, 0.033 mol) was used. Crystalline solid (60 mg, 90% yield): IR ν/cm^-1 3686-3002 (amide and alcohol), 1636 and 1549 (amide), 1513 (Aryl); ¹H NMR (DMSO-d₆, 400 MHz) d 7.92-7.86 (m, NH's), 6.96 (d, 4H, J_11,12 8.2 Hz, H-12), 6.81 (d, 4H, J_9,11 1.6 Hz, H-9), 6.66 (dd, 4H, J_9,11 1.6 Hz and J_11,12 8.2 Hz, H-11), 5.10-4.40 (m, OH's), 4.78 (d, 4H, J_1,2 7.6 Hz, H-1), 3.73 (s, 12H, OCH₃), 3.69 (d, 4H, J_3,4 3.0 Hz, H-4), 3.56 (m, 4H, H-2), 3.52-3.42 (m, 12H, H-5, H-6 and H-6'), 3.38 (dd, 4H, J_2,3 9.6 Hz and J_3,4 3.0 Hz, H-3), 3.07 (bs, 16H, Ha and Hb), 2.74 (t, 8H, H-13, J_13,14 7.8 Hz), 2.60 (t, 8H, J_c,d 7.0 Hz, Hd), 2.36-2.32 (m, 12H, He and H-14), 2.17 (t, 8H, J_c,d 7.0 Hz, Hc), 1.33 (bs, 4H, Hf), 1.22 (bs, 16H, Hg, Hh, Hi and Hj); ¹³C NMR (DMSO-d₆; 100 MHz) δ 171.5 (NHC=O's); 148.8 (C-8); 144.8 (C-7); 135.0 (C-10); 119.9 (C-11); 115.5 (C-12); 112.8 (C-9); 100.9 (C-1); 75.3 (C-5); 73.5 (C-3); 70.3 (C-2); 68.0 (C-4); 60.2 (C-6); 55.6 (OCH₃); 52.8 (Ce); 49.4 (Cd); 38.3 (Ca and Cb); 37.3 (C-14); 33.2 (Cc); 30.7 (C-13); 29.1 ; 27.0 (Cg, Ch, Ci and Cj); 26.6 (Cf); MS-ESI m/z calcd. for C₉₆H₁₄₈N₁₀O₃₆ 2018.3, found 1010.4 (M + 2H⁺).

Pentaerythrityltetramine tetrahydrochloride 12

A mixture of pentaerythritol (6.8 g, 50 mmol) and p-toluenesulfonyl chloride (43 g, 0.226 mol) in dry pyridine (75 mL) was stirred for 48 h under nitrogen atmosphere at room temperature. This mixture was added to 500 mL of 6 mol L^-1 aqueous HCl. The resulting white precipitate was filtered and washed with methanol (300 mL) to afford pentaerythritol tetratosylate (34 g, 90% yield) which was used without further purification.

A mixture of pentaerythritol tetratosylate 11 (3.0 g, 4.0 mmol), sodium azide (3.0 g, 48 mmol) in DMF (30 mL) was stirred for 20 h at 100 ºC. This mixture was added to 200 mL of water and extracted with diethyl ether (4 × 40 mL). The combined organic layers were dried with anhydrous sodium sulfate, filtered and concentrated to about 30 mL. Dioxane (15 mL) was added and the solution was concentrated again to about 30 mL. Dioxane (15 mL), triphenylphosphine (6.3 g, 24 mmol) and 28-30% (w/w) aqueous ammonium hydroxide (22 mL) were added and the mixture was stirred at room temperature for 48 h. The solvent was removed by air jet and chloroform (60 mL) was added to the resulting white solid. The mixture was extracted with 3 mol L^-1 aqueous HCl (5 × 15 mL). The combined aqueous layers were concentrated to about 15 mL and 37% (w/w) aqueous HCl (2 mL) was added. The resulting suspension was filtered and washed with ethanol and diethyl ether. Compound 12 was obtained as an amorphous white solid (0.76 g, 62% yield, 3 steps): IR ν/cm^-1 3124 and 1582 (amine), 2988 and 2863 (aliphatic); ¹H NMR (D₂O, 200 MHz) d 3.43 (s, 8H, CH₂); ¹³C NMR (D₂O; 50 MHz) d 38.6 (CH₂).

Supplementary Information

Supplementary data are available free of charge at http://jbcs.sbq.org.br, as PDF file.

Acknowledgments

We are thankful to Fundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG), Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) for financial support and fellowships and to Professor Renata Barbosa de Oliveira for providing Sephadex LH-20.

Submitted: December 14, 2011

Published online: May 10, 2012

Supplementary Information

Erratum

An Efficient Synthesis of D-Galactose-Based Multivalent Neoglycoconjugates

Saulo F. de Andrade^I; Rute C. Figueiredo^II;José D. de Souza Filhoa and Ricardo J. Alves^*,II

^IDepartamento de Química, Instituto de Ciências Exatas

^IIFaculdade de Farmácia, Universidade Federal de Minas Gerais, Av. Antônio Carlos, 6627, 31270-901 Belo Horizonte-MG, Brazil

Vol. 23, No. 6, 1062-1069, 2012.

http://dx.doi.org/10.1590/S0103-50532012000600010

Page 1062, line 3: Second author address change

Saulo F. de Andrade^I; Rute C. Figueiredo^III; José D. de Souza Filhoa and Ricardo J. Alves^*,II

^IDepartamento de Química, Instituto de Ciências Exatas

^IIFaculdade de Farmácia, Universidade Federal de Minas Gerais, Av. Antônio Carlos, 6627, 31270-901 Belo Horizonte-MG, Brazil

^IIIUniversidade Federal de Ouro Preto, Campus Morro do Cruzeiro, 35400-000 Ouro Preto-MG, Brazil

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Publication Dates

• Publication in this collection
  07 Jan 2013
• Date of issue
  June 2012

History

• Received
  14 Dec 2011
• Accepted
  10 May 2012