J. Orwenyo et al. / Bioorg. Med. Chem. 21 (2013) 4768–4777
4775
0
10 min. Cu(OTf)2 (4 mg, 11
was stirred for 6 h at rt under argon. The reaction was quenched by
addition of triethylamine (20 l) and MeOH (100 l). The solvent
l
mol) was then added and the mixture
4.69/62.68 (Man2 -6a), 4.65/73.43 (Bn-CH2), 4.51/70.69 (Bn-CH2),
4.48/74.33 (Bn-CH2), 4.46/62.68 (Man2-6b), 4.39/69.07, 4.35/
74.71, 4.31/79.38, 4.23/66.43 (Man1-6a), 4.20/55.63 (GlcN-2),
4.08/75.62, 4.07/66.43 (Man1-6b), 4.01/80, 3.98/68.44 (GlcN-6a),
3.94/68.44 (GlcN-6b), 3.60/74.84, 3.42/74.7; MALDI TOF-MS: calcd
for C116H100FNO29 M = 1989.64; found (m/z), 2012.26 [M+Na]+.
l
l
was removed and the syrup was subject to silica gel column chro-
matography (hexane/EtOAc, 3:2) to give the alcohol 12 as a solid
(15 mg, 60%). 1H NMR (500 MHz, CDCl3), d 8.17–6.71 (m, 44H, Ar),
5.87 (t, 1H, J = 10 Hz), 5.70–5.65 (m, 3 H), 5.36–5.31 (m, 1H), 5.03
(d, 1H, J = 8.5 Hz), 4.91 (d, 1H, J = 10.5 Hz), 4.80–4.72 (m, 5H),
4.66–4.63 (m, 2H), 4.53–4.45 (m, 2H), 4.25 (d, 1H, J = 12.5 Hz),
4.20–4.09 (m, 3H), 3.93 (t, 1H, J = 9.5 Hz), 3.85–3.79 (m, 4H),
3.61–3.57 (m, 1H), 3.52 (d, 1H, J = 10 Hz), 3.43 (m, 1H), 3.21 (m,
1H); 1H–13C HSQC NMR (1H at 500 MHz, 13C at 125 MHz, CDCl3) d
8.17/130.11 (Ar), 8.05/130.04 (Ar), 7.92/130.00 (Ar), 7.77/129.71
(Ar), 7.63/133.44 (Ar), 7.61/133.43 (Ar), 7.60/133.44 (Ar), 7.58/
133.43 (Ar), 7.56/133.41 (Ar), 7.54/133.43 (Ar), 7.53/133.56 (Ar),
7.50/128.60 (Ar), 7.47/128.68 (Ar), 7.41/128.11 (Ar), 7.40/133.12
(Ar), 7.40/128.10 (Ar), 7.39/128.44 (Ar), 7.37/128.35 (Ar), 7.36/
128.13 (Ar), 7.27/128.43 (Ar), 7.23/128.13 (Ar), 7.06/127.51 (Ar),
4.10. Synthesis of benzyl 6-deoxy-6-fluoro-
mannopyranosyl-(1?3)-[ -mannopyranosyl-(1?6)]-4-O-
benzyl-b- -mannopyranosyl-(1?4)-2-deoxy-2-acetamido-3,6-
di-O-benzyl-glucopyranoside (15)
a-D-
a-D
D
A solurion of 14 (15 mg, 7.5
containing hydrazine monohydrate (600
7 h. The solvent was then removed and the residue was dissolved
in MeOH (5 ml) and 1.0 M aqueous NaHCO3 (5 ml). Acetic anhy-
lmol) in 90% aqueous EtOH (4 ml)
l
l) was stirred at 80 °C for
dride (30 ll) was added and the mixture was stirred at rt for 1 h.
The solution was concentrated and the residue was subject to pre-
parative HPLC purification to give 15 as a white powder (5 mg, 60%
for two steps). 1H NMR (500 MHz, D2O) d 7.53–7.34 (m, 20H, Ar),
5.14 (s, 1H), 5.08 (s, 1H), 4.94–4.89 (m, 3H), 4.84–4.79 (m, 3H),
4.75–4.63 (m, 6H), 4.18–4.1 (m, 4H), 4.0 (br s, 1H), 3.97–3.9 (m,
5H), 3.88–3.85 (m, 3H), 3.82–3.76 (m, 3H), 3.74–3.71 (m, 2H),
3.66–3.61 (m, 4H), 1.9 (s, 3H, CH3); 1H–13C HSQC NMR (1H at
500 MHz, 13C at 125 MHz, D2O) d 7.53/127.82 (Ar), 7.49/128.21
(Ar), 7.48/128.14 (Ar), 7.47/127.92 (Ar), 7.42/127.84 (Ar), 7.41/
7.01/127.82 (Ar), 6.83/127.08 (Ar), 6.78/127.82 (Ar), 6.71/127.21
0
(Ar), 5.87/65.94, 5.70/70.00 (Man2 -2), 5.69/65.94, 5.65/71.88
0
(Man1-2), 5.36/99.69 (Man2 -1), 5.03/97.34 (GlcN-1), 4.91/75.73
(½ Bn), 4.80/98.36 (Man1-1), 4.79/73.63 (Bn-CH2), 4.76/74.20 (Bn-
1
CH2), 4.76/70.86 (Bn-CH2), 4.72/81.73 (C-6a, JC–F = 195 Hz), 4.66/
75.73 (Bn-CH2), 4.63/81.73 (C-6b), 4.53/73.63 (Bn-CH2), 4.45/
70.86 (Bn-CH2), 4.25/74.20 (Bn-CH2), 4.20/55.64 (GlcN-2), 4.17/
78.75, 4.09/76.37, 3.93/74.70, 3.85/79.98, 3.82/68.17 (GlcN-6a),
3.82/61.97 (Man1-6a), 3.79/68.17 (GlcN-6b), 3.60/61.97 (Man1-
6b), 3.52/74.38, 3.43/70.78, 3.21/75.52; 19F NMR (376 MHz, CDCl3),
127.72, 7.36/127.19, 5.14/103.13 (Man2-1), 5.08/73.61 (Bn-CH2),
0
4.94/70.47 (Bn-CH2), 4.91/100.00 (Man2 -1), 4.90/74.59 (Bn-CH2),
2
3
1
ꢂ232.2 (td, JF–H = 47.1 Hz, JF–H = 24.0 Hz). MALDI TOF-MS: calcd
4.84/73.61 (Bn-CH2), 4.81/82.81 (Man2-6a, JC–F = 160 Hz), 4.79/
for C82H74FNO20 M = 1411.47; found (m/z), 1434.56 [M+Na]+.
73.13 (Bn-CH2), 4.74/82.81(Man2-6b), 4.74/74.59 (Bn-CH2), 4.69/
99.93(Man1-1), 4.68/70.47 (Bn-CH2), 4.65/100.31 (GlcNAc-1),
4.63/73.13 (Bn-CH2), 4.19/72.53, 4.16/70.84, 4.15/72.49 (Man1-2),
4.9. Synthesis of benzyl 2,3,4-tri-O-benzoyl-6-deoxy-6-fluoro-
a-
D
-mannopyranosyl-(1?3)-[2,3,4,6-tetra-O-benzoyl-
a-D
-
4.11/77.10, 4.0/70.95 (Man2-2), 3.96/71.09, 3.93/54.99 (GlcNAc-
0
mannopyranosyl-(1?6)]-2-O-benzoyl-4-O-benzyl-b-
D-
2), 3.93/68.45 (GlcNAc-6a), 3.92/70.63 (Man2 -2), 3.90/68.45 (Glc-
mannopyranosyl-(1?4)-3,6-di-O-benzyl-2-deoxy-2-
phthalimido-b-D-glucopyranoside (14)
NAc-6b), 3.87/74.06, 3.85/71.27, 3.85/65.83 (Man-6a), 3.81/79.69,
3.79/66.24, 3.75/67.18, 3.73/61.24 (Man-6a), 3.71/65.83 (Man-
6b), 3.68/61.24 (Man-6b), 3.64/82.82, 3.62/73.108, 3.61/74.65,
2
A mixture of 12 (15 mg, 10.6
4 Å molecular sieves (200 mg) in CH2Cl2 (2 ml) was stirred at rt un-
der argon for 10 min, then TMSOTf (1.5 l, 8 mol) was added, and
the resulting mixture was stirred at rt for 16 h. The reaction was
quenched by addition of triethylamine (100 l) and the solution
l
mol), 13 (30 mg, 40.5
l
mol), and
1.90/21.67 (CH3); 19F NMR (376 MHz, D2O), d ꢂ233.4 (td, JF–
H = 47.6 Hz, 3JF–H = 24 Hz). ESI-MS: calcd for C54H68FNO20
M = 1069.43; found (m/z), 1070.74 [M+H]+.
,
l
l
l
4.11. Synthesis of 6-deoxy-6-fluoro-
(1?3)-[ -mannopyranosyl-(1?6)]-b-
(1?4)-2-deoxy-2-acetamido- -glucopyranose (16)
a-
D
-mannopyranosyl-
was filtered through a Celite pad. The filtrate was concentrated
and the residue was subject to silica gel column chromatography
(hexanes/EtOAc, 2:1 to 1:1) to afford 14 as a white solid (16 mg,
73%). 1H NMR (500 MHz, CDCl3), d 8.39–6.44 (m, 64H Ar), 6.18 (t,
1H, J = 10 Hz), 6.05–6.0 (m, 2H), 5.92–5.86 (m, 4H), 5.50 (s, 1H),
5.32–5.30 (m, 2H), 5.09 (d, 1H, J = 8.5 Hz), 4.98–4.89 (m, 2H), 4.94
(t, 1H, J = 11 Hz), 4.85–4.76 (m, 4H), 4.75–4.65 (m, 3H), 4.51–4.46
(m, 3H), 4.39–4.31 (m, 3H), 4.23–4.20 (m, 2H), 4.08–4.07 (m, 2H),
4.01–3.98 (m, 2H), 3.94–3.92 (m, 1H), 3.60 (d, 1H, J = 9.5 Hz), 3.42
(d, 1H, J = 9.5 Hz); 1H–13C HSQC NMR (1H at 500 MHz, 13C at
125 MHz, CDCl3) d 8.39/130.21 (Ar), 8.20/129.94 (Ar), 8.11/129.99
(Ar), 8.08/128.88 (Ar), 8.0/129.77 (Ar), 7.86/129.70 (Ar), 7.74/
133.44 (Ar), 7.73/122.98 (Ar), 7.67/133.12 (Ar), 7.61/128.73 (Ar),
7.60/127.68 (Ar), 7.58/133.13 (Ar), 7.54/128.03 (Ar), 7.52/133.11
(Ar), 7.51/128.44 (Ar), 7.40/128.43 (Ar), 7.37/122.89 (Ar), 7.31/
128.14 (Ar), 7.14/127.49 (Ar), 7.08/128.03 (Ar), 7.07/127.74 (Ar),
6.57/126.57 (Ar), 6.56/128.20 (Ar), 6.44/127.44 (Ar), 6.18/66.57,
6.05/65.94, 6.0/70.32, 5.92/69.72, 5.91/69.82 (Man2-2), 5.87/71.88
a-
D
D-mannopyranosyl-
D
A mixture of 15 (5 mg, 4.7 lmol) and palladium–carbon (5%,
25 mg) in MeOH (2 ml) was stirred for 16 h under a H2 atmosphere.
Then the mixture was filtered through a Celite pad and the filtrate
was concentrated to dryness. The residue was dissolved in water
and lyophilized to give free tetrasaccharide 16 as a white solid
(3 mg, 91%). 1H NMR (500 MHz, D2O) d 5.27 (d, 1H, J = 3 Hz), 5.18
(s, 1H), 4.96 (s, 1H), 4.88–4.78 (br s, 3H), 4.71–4.74 (m, 1H), 4.27–
4.24 (m, 2H), 4.16–4.14 (m, 2H), 4.07–4.06 (m, 2H), 4.03–4.0 (m,
3H), 3.98–3.88 (m, 10H), 3.85–3.77 (m, 8H), 3.72–3.67 (m, 4H),
3.63–3.58 (m, 1H), 2.11 (s, 3H, CH3); 1H–13C HSQC NMR (1H at
500 MHz, 13C at 125 MHz, D2O) d 5.27/90.53, 5.18/102.68 (Man2-
0
1), 4.96/99.66 (Man2 -1), 4.85/100.34 (Man1-1), 4.78/94.98 (Glc-
1
NAc-1), 4.83/82.6 (Man2-6a, JC–F = 168 Hz), 4.73/82.6 (Man2-6b),
4.28/70.35, 4.24/70.31(Man1-2), 4.16/68.48, 4.15/69.98 (Man2-2),
0
4.07/72.18, 4.06/70.0, 4.03/69.98 (Man2 -2), 4.02/71.90, 4.0/69.12,
0
(Man1-2), 5.86/70.30 (Man2 -2), 5.50/99.70 (Man2-1, JC-1/H-
3.99/69.12, 3.98/70.1, 3.98/65.93 (Man1-6a), 3.97/69.13, 3.96/
70.31, 3.95/60.97 (6a), 3.95/53.68, 3.94/70.34, 3.91/60.31 (6a),
3.88/65.93(Man1-6b), 3.85/60.31(6b), 3.84/72.15, 3.83/80.94, 3.82/
65.63, 3.81/60.97 (6b), 3.78/72.48, 3.78/55.95 (GlcNAc-2), 3.77/
80.03, 3.72/74.10, 3.71/72.79, 3.70/66.87, 3.68/66.88, 3.61/74.38,
1
0
1
1 = 173 Hz,
a
), 5.32/98.28 (Man2 -1, JC-1/H-1 = 176 Hz,
a
), 5.30/
1
75.63 (Bn-CH2), 5.09/97.20 (GlcN-1, JC-1/H-1 = 163 Hz, b), 4.98/
99.07 (Man1-1, JC-1/H-1 = 162 Hz, b), 4.99/75.63 (Bn-CH2), 4.94/
1
73.43 (Bn-CH2), 4.85/81.72(Man2-6a, JC–F = 195 Hz), 4.81/70.69
1
(Bn-CH2), 4.78/74.33 (Bn-CH2), 4.76/81.72 (Man2-6b), 4.75/70.32,
2.12/22.12 (CH3); 19F NMR (376 MHz, D2O), d -233.6 (td, JF–
2