Katajisto et al.
The residue was purified by column chromatography, eluting
first with a 1:6 (v/v) mixture of EtOAc and toluene and then
with a 6:5 (v/v) mixture of EtOAc in petroleum ether. The yield
Syn th esis of Glycoclu ster s 21-24. The Fmoc group was
removed from support 17, as described above. The support was
washed with DCM and MeOH and dried under reduced
pressure. The support (50 to 150 mg) was then suspended in
1 mL of DMF, and the branching building block 1 (3 molar
equiv in 0.5 mL) was added together with HATU (3 molar
equiv in 0.5 mL) and DIPEA (6 molar equiv). The mixture was
shaken for 2 h at room temperature, then the support was
filtered, washed with DMF, DCM, and MeOH, and dried under
reduced pressure. The Fmoc group was removed, and the
support was again washed with DCM and MeOH and dried
under reduced pressure. The first O-(glycosyl)-N-Fmoc-L-Ser-
OPfp building block (3 molar equiv in 0.5 mL of DMF) and
HOBt (7.5 molar equiv in 0.5 mL of DMF) were added onto
the support suspended in 1 mL of DMF. After 3 h of shaking
at room temperature, the support was filtered, washed with
DMF, DCM, and MeOH, and dried under reduced pressure.
The Boc group was removed from the branching unit with 25%
TFA in DCM (1 h at room temperature), and the filtered
support was washed with 10% pyridine in DCM and then
consecutively with DMF, DCM, and MeOH and dried under
reduced pressure. The support was suspended in 1 mL of DMF,
and the second glycosylated N-Fmoc-L-Ser-OPfp building block
(4 molar equiv in 0.5 mL of DMF) and HOBt (10.0 molar equiv
in 0.5 mL of DMF) were added. After 3 h of shaking at room
temperature, the support was filtered, washed with DMF,
DCM, and MeOH, and dried under reduced pressure. Finally,
the Alloc group was removed via a palladium-catalyzed hy-
drostannolysis by Bu3SnH.18 Accordingly, a mixture of Pd-
(OAc)2 (2 molar equiv) and PPh3 (12 molar equiv) in DMF (1
to1.5 mL) and AcOH (12 molar equiv) was added onto the
support (50-150 mg). After this, Bu3SnH (12 molar equiv) was
added to the reaction mixture. The mixture was shaken at
room temperature for 15 min. The resin was filtered, washed
with 10% pyridine in DCM and then with DMF, DCM, and
MeOH, and dried under reduced pressure. The last glycosyl-
ated Fmoc-Ser-OPfp building block (7 molar equiv in 0.5 mL
of DMF) and HOBt (17.5 molar equiv in 0.5 mL of DMF) were
added onto the support suspended in 1 mL of DMF. The
mixture was shaken for 5 h at room temperature. The support
was filtered, washed with DMF, DCM, and MeOH, and dried
under reduced pressure. After each coupling step, the product
was released from an aliquot of the solid support and analyzed
by HPLC. Only minor impurities gradually appeared. The
coupling efficiency of the synthesis was 80-90%.
1
of the pure â-anomer was 15%. H NMR (CDCl3, 400 MHz) δ
7.77 (d, 2H, J ) 7.0 Hz), 7.61 (d, 2H, J ) 7.0 Hz), 7.41 (t, 2H,
J ) 7.0 Hz), 7.32 (t, 2H, J ) 7.5 Hz), 5.69 (d, 1H, J ) 8.5 Hz),
5.23 (t, 1H, J ) 9.4 Hz), 5.09 (t, 1H, J ) 9.7 Hz), 4.98 (dd, AB
type, 1H, J ) 9.4 and 8.1 Hz), 4.87 (m, 1H), 4.54 (m, 2H), 4.43
(m, 2H), 4.11-4.26 (m, 3H), 3.99 (dd, 1H, J ) 3.5 and 10.5
Hz), 3.68 (m, 1H), 2.05, 2.03, 1.98 (3s, 12H); 13C NMR (CDCl3,
100 MHz) δ 170.7, 170.6, 170.2, 169.4, 169.3, 166.1 155.8,
143.6, 143.5, 141.4, 141.3, 127.8, 127.1, 125.0, 124.9, 120.1,
100.7, 72.5, 72.1, 71.1, 69.8, 68.2, 67.3, 61.7, 54.2, 47.1, 20.6,
20.5; HRMS (ESI) [M
824.1996.
+
H]+ calcd 824.1972, obsd
N-(9-F lu or en ylm eth oxyca r bon yl)-3-O-(2,3,4,6-tetr a -O-
a cetyl-r-D-m a n n op yr a n osyl)-L-ser in e P en ta flu or op h en yl
Ester (15):20 Compound 15 was prepared as described for 14,
using 1,2,3,4,5-penta-O-acetyl-R-D-mannopyranose (1 g, 2.6
mmol) as a starting material. Compound 15 was obtained as
1
white amorphous solid in a 35% yield. H NMR (CDCl3, 400
MHz) δ 7.76 (d, 2H, J ) 7.5 Hz), 7.63 (d, 2H, J ) 6.2 Hz), 7.40
(t, 2H, J ) 7.3 Hz,), 7.31 (t, 2H, J ) 7.3 Hz), 6.03 (d, 1H, J )
8.5 Hz), 5.24-5.33 (m, 2H), 4.99 (m, 1H), 4.88 (br s, 1H), 4.48
(m, 3H), 4.12-4.28 (m, 4H), 4.08 (dd, 1H), 3.99 (m, 1H), 2.01,
2.04, 2.06, 2.17 (4s, 12H); 13C NMR (CDCl3, 100 MHz) δ 170.5,
169.8, 169.7, 166.2, 155.7, 143.5, 141.3, 127.8, 127.1, 125.0,
120.0, 99.0, 69.9, 69.4, 69.1, 68.6, 67.6, 65.9, 62.4, 54.4, 47.0,
21.4, 20.6, 20.8; HRMS (ESI) [M + H]+ calcd 824.1972, obsd
824.2007.
N-(9-Flu or en ylm eth oxycar bon yl)-3-O-(2,3,4-tr i-O-acetyl-
â-D-r ibopyr an osyl)-L-ser in e P en taflu or oph en yl Ester (16).
Compound 16 was prepared as described for 14, using 1,2,3,4-
tetra-O-acetyl-â-D-ribopyranose (500 mg, 1.6 mmol) as a start-
ing material. The product was obtained as white amorphous
solid in a 38% yield. 1H NMR (CDCl3, 400 MHz) δ 7.76 (d, 2H,
J ) 7.5 Hz), 7.62 (d, 2H, J ) 7.5 Hz), 7.40 (t, 2H, J ) 7.5 Hz),
7.32 (t, 2H, J ) 7.5 Hz), 5.79 (d, 1H, J ) 8.9 Hz), 5.36 (t, 1H,
J ) 3.4 Hz), 5.14 (m 1H), 5.08 (t, 1H, J ) 3.2 Hz), 4.96 (m,
1H), 4.79 (d, 1H, J ) 3.6 Hz), 4.38-4.57 (m, 3H), 4.26 (t, 1H),
3.94-3.80 (m, 3H), 2.12, 2.11, 2.08 (3s, 9H); 13C NMR (CDCl3,
100 MHz) δ 171.1, 169.9, 169.7, 169.5, 166.3, 155.8, 143.6,
143.5, 141.3, 127.8, 127.1, 125.0, 120.0, 98.8, 68.3, 67.9, 67.6,
66.4, 65.8, 54.0, 47.0, 20.8, 20.7, 20.6; HRMS (ESI) [M + H]+
calcd 752.1761, obsd 752.1736.
Clea va ge fr om Solid Su p p or t. The solid support (50 to150
mg) was first swollen with a small amount of DCM and then
1 mL of 0.1% HBr in AcOH was added. The mixture was
shaken for 2 h at room temperature. The support was filtered,
washed with DCM and MeOH, and dried under reduced
pressure. The support was suspended in a mixture of TFA and
DCM (1 mL, 1:1, v/v). After 2 h of shaking at room tempera-
ture, the solution was collected by filtration and evaporated
under reduced pressure. After HPLC purification, the authen-
ticity of 21-24 was verified by HPLC/ESI-MS (Table 1). In
addition, glycocluster 21 prepared in a larger quantity was
characterized by HRMS and 1H and 13C NMR spectroscopy
(including PDQF and HMBC spectra).
Solid Su p p or t 17. Aminomethyl polystyrene that was
employed as a solid support was first acylated with a Fmoc-
protected SCAL linker16 using standard HATU activation.17
Accordingly, 150 mg of the solid support was suspended in 1
mL of DMF, Fmoc-SCAL (2 molar equiv, 19.3 mg in 0.5 mL of
DMF), HATU (3 molar equiv, 17.1 mg in 0.5 mL of DMF), and
diisopropylethylamine (DIPEA, 4 molar equiv, 10.5 µL) were
added, and the mixture was shaken for 3 h at room temper-
ature. The support was filtered and washed with DMF, DCM,
and MeOH, and finally dried under reduced pressure. The
nonreacted amino groups were capped with Ac2O in THF
containing N-methylimidazole and lutidine. The Fmoc groups
were removed with 25% piperidine in DMF (15 min at room
temperature). According to UV spectroscopic quantification of
piperidinyl benzofulvene released, the loading was 120 µmol
g-1. The exposed amino function of the SCAL linker was then
acylated with N-Fmoc-glycine by adding N-Fmoc-glysine (5
molar equiv, 30 mg in 0.5 mL of DMF), HATU (5 molar equiv,
38 mg in 0.5 mL of DMF), and DIPEA (10 molar equiv, 35 µL)
to a suspension of the support in DMF (1 mL), and shaking
the mixture for 1 h at room temperature. The solid support
was filtered, washed with DMF, DCM, and MeOH, and dried
under reduced pressure to afford support 17.
Glycoclu ster 21. Starting from 150 mg of solid support,
1
the overall isolated yield was 3.8 mg (10%). H NMR (DMSO-
d6, 90 °C, 500 MHz) δ 7.82 (d, 6H, J ) 7.6 Hz), 7.63 (t, 6H, J
) 7.2 Hz), 7.38 (t, 6H, J ) 7.3 Hz), 7.30 (t, 6H, J ) 7.3 Hz),
5.25 (d, 2H, J ) 3.0 Hz), 5.18 (t, 1H, J ) 9.0 Hz), 5.09 (dd, 2H,
J ) 10.5 and 3.5 Hz), 4.88-4.95 (m, 4H), 4.71-4.79 (m, 3H),
4.64 (d, 2H, J ) 7.3 Hz), 4.18-4.30 (m, 14H), 4.15 (m, 1H),
4.09 (t, 2H, J ) 6.8 Hz), 4.01-4.04 (m, 6H), 3.78-3.90 (m, 8H),
3.72 (dd, 1H, J ) 13.0 and 5.3 Hz), 3.62 (dd, 1H, J ) 16.1 and
5.3 Hz), 1.98, 1.96, 1.92, 1.91, 1.89 (5s, 36H); 13C NMR (DMSO-
d6, 90 °C, 100 MHz) δ 171.4, 171.2, 169.6, 169.5, 169.4, 169.3,
168.9, 168.8, 168.6, 168.5, 168.4, 143.5, 143.4, 140.3, 140.2,
127.1, 126.6, 124.7, 124.6, 119.5, 99.6, 99.2, 72.0, 70.6, 70.5,
70.0, 69.8, 68.3, 68.2, 68.1, 67.0, 65.8, 61.4, 60.7, 54.8, 54.7,
(20) For previous synthesis of 15, see: Kragol, G.; Otvos, L, J r.
Tetrahedron 2001, 57, 957.
8000 J . Org. Chem., Vol. 67, No. 23, 2002