Synthesis of Oligonucleotides Containing Fapy‚dG
NMR (CDCl3) δ 10.31 (d, 0.7 H, J ) 8.8 Hz), 9.78 (d, 0.3 H, J
) 8.4 Hz), 7.17-7.41 (m, 13 H), 6.79-6.91 (m, 4 H), 6.32 (t,
0.7 H, J ) 8.0 Hz), 6.17 (dd, 0.3 H, J ) 8.0, 13.2 Hz), 4.67 (s,
0.6 H), 4.63 (s, 1.4 H), 4.40 (d, 0.7 H, J ) 4.8 Hz), 4.30 (m, 0.3
H), 4.24 (dd, 0.7 H, J ) 3.6, 5.2 Hz), 4.00 (m, 0.3 H), 3.76 (s,
6 H), 3.13 (m, 2 H), 2.89 (septet, 1 H, J ) 6.8 Hz), 2.35 (m, 1
H), 2.00 (d, 1 H, J ) 13.6 Hz), 1.22 (m, 6 H), 0.89 (m, 6.3 H),
154.7, 154.6, 154.5, 150.6, 149.6, 149.4, 144.0, 143.9, 143.4,
143.3, 135.0, 134.95, 134.90, 130.0, 128.11, 128.05, 128.0,
127.7, 127.3, 114.6, 113.63, 113.55, 113.3, 111.74, 111.70,
87.21, 87.16, 86.0, 85.93, 85.86, 84.5, 84.4, 84.3, 84.23, 84.15,
82.6, 82.5, 78.1, 77.8, 73.7, 67.6, 67.4, 67.2, 62.8, 62.5, 55.2,
54.7, 54.61, 54.58, 54.5, 44.5, 44.4, 40.6, 39.1, 33.2, 25.9 24.1,
23.6, 22.9, 22.6, 18.3, 11.7, 11.6, -4.7, -4.8; 31P NMR (CDCl3)
δ -0.06, -0.042; IR (KBr film) 3248, 2958, 1686, 1624, 1584,
1510, 1421, 1251, 1179, 1031, 840, 789, 558 cm-1; HRMS (M
+ Na) calcd for C58H72N7O17NaSiP 1220.4384, found 1220.4328.
Preparation of 8b. The reaction between 7b (252 mg, 0.44
mmol) and 5′-DMT-dT-O-methyl phosphoramidite (368 mg,
0.525 mmol) was carried out as described above for 7a using
tetrazole (54 mg, 1.71 mL of 0.45 M, 0.77 mmol), followed by
t-BuOOH (0.30 mL of 5-6 M solution in decane, 1.50 mmol).
The reaction mixture was concentrated and chromatographed
(2% methanol in CH2Cl2) to give product 8b (250 mg, 47%):
1H NMR (CD3CN) δ 9.54 (d, 1 H, J ) 8.0 Hz), 9.42 (bd s, 1 H),
7.15-7.44 (m, 14 H), 6.80-6.91 (m, 6 H), 6.17-6.28 (m, 2 H),
5.08-5.18 (m, 1 H), 4.72 (t, 2 H, J ) 4.0 Hz), 4.42 (m, 1 H),
4.15-4.21 (m, 1 H), 4.03-4.10 (m 1 H), 3.93 (m, 1 H), 3.79 (m,
1 H), 3.73-3.74 (m, 6 H), 3.63-3.70 (m, 3 H), 3.29-3.34 (m, 2
H), 2.84 (septet, 1 H, J ) 6.8 Hz), 2.40-2.55 (m, 2 H), 2.16-
2.23 (m, 2 H), 1.47 (t, 1.5 H, J ) 1.2 Hz), 1.47 (dd, 1.5 H, J )
1.2, 4.0 Hz), 1.18 (m, 6 H), 0.88 (d, 9 H, J ) 1.2 Hz), 0.08 (m,
6 H); 13C NMR (CD3CN) δ 173.0, 164.7, 159.8, 156.2, 151.4,
151.2, 145.73, 145.68, 143.6, 136.6, 136.4, 131.0, 129.0, 128.4,
128.3, 128.0, 115.6, 115.2, 114.6, 114.1, 111.7, 111.6, 87.7, 85.8,
85.1, 85.0, 83.2, 79.1, 76.7, 73.2, 68.2, 68.0, 65.8, 64.1, 63.92,
63.85, 55.9, 55.5, 55.41, 55.35, 52.6, 42.0, 39.2, 34.0, 30.9, 26.1,
24.4, 18.6, 12.2, -4.5, -4.7; 31P NMR (CD3CN) δ 8.45, -1.64;
IR (KBr film) 3224, 2954, 1690, 1608, 1584, 1508, 1472, 1249,
1179, 1032, 840, 832, 785 cm-1; HRMS (M + Na) calcd for
C58H72N7O17NaSiP 1220.4384, found 1220.4418.
Preparation of 9 from 8a. Compound 8a (1.0 g, 0.84
mmol) was reduced by H2 (80 psi) in the presence of palladium
on carbon (10%, 728 mg) and diisopropylethylamine (0.89 g,
1.2 mL, 6.8 mmol) in THF (40 mL) for 2 h at 25 °C. The
reaction mixture was cooled to 0 °C, whereupon acetic formic
anhydride (0.220 mL, 2.51 mmol) and pyridine (268 mg, 0.274
mL, 3.39 mmol) were added. After 10 min at 0 °C, the reaction
mixture was filtered through Celite. After concentration, the
residue was chromatographed (3-5% methanol in CH2Cl2) to
afford 9a (less polar, 431 mg, 43%) and 9b (110 mg, 11%). 9a:
1H NMR (CD3OD) δ 8.20 (s, 0.3 H), 8.19 (s, 0.3 H), 7.87 (s, 0.1
H), 7.76 (s, 0.1 H), 7.60 (s, 1 H), 7.08-7.39 (m, 12 H), 6.82-
6.91 (m, 6 H), 6.33 (m, 1 H), 6.10-6.14 (m, 1 H), 5.13 (m, 1
H), 4.65-4.68 (m, 2 H), 4.34 (m, 2 H), 3.99-4.08 (m, 3 H),
3.70-3.79 (m, 9 H), 3.44 (dd, 2 H, J ) 3.2, 11.6 Hz), 2.83
(septet, 1 H, J ) 6.8 Hz), 2.51-2.64 (m, 2 H), 2.38 (m, 1 H),
1.92-1.97 (m, 1 H), 1.39 (d, 1.5 H, J ) 1.2 Hz), 1.38 (d, 1.5 H,
J ) 1.2 Hz), 1.19 (d, 6 H, J ) 6.8 Hz), 0.91-0.93 (m, 9 H),
0.12-0.15 (m, 6 H); 13C NMR (CD3OD) δ 210.0, 173.0, 169.1,
166.0, 163.9, 163.8, 160.3, 159.2, 159.0, 157.0, 156.9, 152.2,
149.5, 145.8, 143.7, 137.3, 137.2, 136.6, 136.5, 131.4, 129.4,
129.0, 128.4, 128.2, 115.7, 114.3, 112.1, 94.5, 88.4, 86.0, 85.9,
85.8, 85.1, 83.3, 83.2, 80.0, 74.4, 74.3, 69.1, 68.7, 68.2, 64.3,
55.8, 55.6, 55.5, 55.4, 41.0, 39.8, 34.6, 30.7, 29.5, 26.4, 24.6,
19.0, 12.2, -4.6; 31P NMR (CD3OD) δ -1.85, -2.20; IR (KBr
film) 3401, 3246, 2956, 1696, 1655, 1583, 1510, 1460, 1252,
1179, 1032, 831, 778, 584 cm-1; HRMS (M + Na) calcd for
C59H74N7O16NaSiP 1218.4591, found 1218.4638. 9b: 1H NMR
(CD3OD) δ 8.17 (s, 0.4 H), 8.18 (s, 0.4 H), 7.93 (s, 0.1 H), 7.92
(s, 0.1 H), 7.61 (d, 0.5 H, J ) 1.2 Hz), 7.57 (d, 0.5 H, J ) 1.2
Hz), 7.11-7.40 (m, 12 H), 6.83-6.92 (m, 6 H), 6.28 (m, 1 H),
6.09 (m, 1 H), 5.11 (m, 1 H), 4.68 (s, 2 H), 4.38 (m, 1 H), 4.27
(m, 1 H), 4.08 (m, 1 H), 4.02 (m, 1 H), 3.84-3.88 (m, 1 H),
3.75 (s, 6 H), 3.72 (dd, 3 H, J ) 11.2, 15.2 Hz), 3.40 (m, 2 H),
2.83 (septat, 1 H, J ) 6.8 Hz), 2.47-2.59 (m, 2 H), 2.20 (m, 1
H), 2.08 (m, 1 H), 1.41 (d, 1.5 H, 1.2 Hz), 1.40 (d, 1.5 H, 1.2
Hz), 1.20 (d, 6 H, J ) 6.8 Hz), 0.89 (d, 9 H, J ) 2.0 Hz), 0.08-
0.10 (m, 6 H); 13C NMR (CD3OD) δ 210.0, 173.04, 172.99,
0.86 (m, 2.7 H), 0.09 (s, 4.2 H), 0.01 (d, 1.8 H, J ) 6.0 Hz); 13
C
NMR (CDCl3), δ 170.5, 158.5, 157.8, 154.3, 148.9, 144.6, 143.9,
135.8, 135.7, 130.0, 127.9, 127.8, 126.8, 114.7, 113.1, 87.6, 86.6,
86.4, 82.3, 74.4, 67.2, 63.5, 55.2, 53.8, 40.2, 33.3, 29.3, 26.0,
25.8, 24.1, 18.4, -4.6, -4.7; IR (KBr film) 3248, 2958, 1686,
1623, 1584, 1510, 1420, 1251, 1179, 1031, 840, 789, 558 cm-1
;
HRMS (M + Na) calcd for C47H57N5O10NaSi 902.3767, found
902.3774.
Preparation of 7. A solution of FeCl3‚6H2O (615 mg, 2.28
mmol) in CH3CN (2 mL) was added to 6 in CH3CN (100 mL)
and stirred for 60 min at room temperature. Additional FeCl3‚
6H2O (307 mg, 1.14 mmol) in CH3CN (2 mL) was added, and
the reaction was stirred 30 min longer. The reaction was
quenched by diisopropylethylamine (0.165 g, 0.222 mL, 1.3
mmol) and methanol (0.162 mL). After concentration, the
mixture was purified by column chromatography (1-4%
methanol in CH2Cl2) to afford fast isomer 7a (4.1 g, 72%) and
slow isomer 7b (0.69 g, 12%). The fast isomer was recrystal-
lized from CH3CN (12 mL) to provide long, white needlelike
crystals (2.3 g, mp 145-147 °C). 7a: 1H NMR (CDCl3) δ 10.26
(d, 1 H, J ) 8.4 Hz), 9.57 (bd s, 1 H), 7.19 (d, 2 H, J ) 8.8 Hz),
6.89 (d, 2 H, J ) 8.8 Hz), 6.25 (t, 1 H, J ) 7.6 Hz), 4.64 (s, 2
H), 4.44 (d, 1 H, J ) 5.6 Hz), 4.15 (dd, 1 H, J ) 1.2, 3.6 Hz),
3.67 (dd, 1 H, J ) 4.4, 12 Hz), 3.58 (dd, 1 H, J ) 3.6, 12 Hz),
2.86 (septet, 1 H, J ) 6.8 Hz), 2.34 (m, 1 H), 2.02 (d, 1 H, J )
13‚6 Hz), 1.21 (d, 6 H, J ) 6.8 Hz), 0.89 (s, 9 H), 0.12 (d, 6 H,
J ) 3.6 Hz); 13C NMR (CDCl3) δ 170.9, 158.1, 154.5, 149.2,
143.8, 127.8, 114.9, 113.7, 88.4, 82.5, 73.9, 67.5, 62.8, 41.0, 33.3,
30.9, 25.9, 24.1, 18.3, -4.7, -4.8; IR (KBr film) 3462, 3242,
2957, 1680, 1628, 1582, 1511, 1421, 1244, 1027, 839, 788 cm-1
;
HRMS (M + Na) calcd for C26H39N5O8NaSi 600.2460, found
600.2447. 7b: 1H NMR (CDCl3), slow isomer, δ 9.98 (d, 1 H, J
) 8.4 Hz), 9.20 (bs, 1 H), 7.21 (d, 2 H, J ) 8.8 Hz), 6.91 (d, 2
H, J ) 8.8 Hz), 6.25 (dd, 1 H, J ) 6.4, 14.4 Hz), 4.65 (s, 2 H),
4.48 (m, 1 H), 3.96 (dd, 1 H, J ) 2.8, 5.6 Hz), 3.67-3.82 (m, 2
H), 2.88 (septet, 1 H, J ) 6.8 Hz), 2.17-2.30 (m, 2 H), 1.22 (d,
6 H, J ) 6.8 Hz), 0.88 (s, 9 H), 0.08 (s, 6 H); 13C NMR (CDCl3)
δ 170.7, 158.2, 154.3, 149.2, 143.9, 127.9, 114.8, 113.8, 87.5,
82.3, 72.7, 67.2, 55.7, 43.6, 42.3, 33.3, 25.7, 24.1, 18.6, 18.0,
17.2, 12.4, -4.7, -4.8; IR (KBr film) 3401, 3236, 2958, 1686,
1624, 1511, 1426, 1250, 1098, 842, 788 cm-1; HRMS (M + Na)
calcd for C26H39N5O8NaSi 600.2460, found 600.2487.
Preparation of 8a. A mixture of 7a (600 mg, 1.04 mmol)
and 5′-DMT-dT-O-methyl phosphoramidite (870 mg, 1.24
mmol) was coevaporated with CH3CN (2 × 4 mL). Tetrazole
(128.6 mg, 4.08 mL of 0.45 M in acetonitrile, 1.84 mmol) was
added to the dried material that was dissolved in CH3CN (16
mL) at 0 °C. After 1 h at 25 °C, the solution was cooled to 0
°C, t-BuOOH (0.724 mL of 5-6 M solution in decane, 3.62
mmol) was added, and stirring was continued for 30 min. The
reaction mixture was concentrated in vacuo and chromato-
graphed (2-2.5% methanol in CH2Cl2) to give product 8a
(1.045 g, 84%): 1H NMR (CDCl3) δ 10.27 (d, 0.5 H, J ) 8.4
Hz), 10.23 (d, 0.5 H, J ) 8.4 Hz), 9.58 (s, 0.5 H), 9.44 (s, 0.5
H), 7.60 (d, 0.5 H, J ) 1.2 Hz), 7.57 (d, 0.5 H, J ) 1.2 Hz),
7.11-7.36 (m, 12 H), 6.81-6.84 (m, 6 H), 6.31 (m, 2 H), 5.09-
5.15 (m, 1 H), 4.70 (d, 1 H, J ) 1.6 Hz), 4.66 (d, 1 H, J ) 4.0
Hz), 4.44 (t, 1 H, J ) 5.6 Hz), 4.18-4.32 (m, 2 H), 3.99-4.07
(m, 2 H), 3.78 (s, 6 H), 3.71 (dd, 3 H, J ) 11.6, 23.2 Hz), 3.51-
3.55 (m, 1 H), 3.40 (dd, 1 H, J ) 10.8, 16.0 Hz), 2.84 (septet,
1 H, J ) 6.8 Hz), 2.63 (m, 1 H), 2.48 (m, 1 H), 2.31 (m, 1 H),
1.9 (dd, 1 H, J ) 2.8, 13.2 Hz), 1.35 (d, 1.5 H, J ) 0.4 Hz),
1.33 (d, 1.5 H, J ) 0.8 Hz), 1.20 (d, 6 H, J ) 7.2 Hz), 0.92 (d,
9 H, J ) 2.0 Hz), 0.15 (s, 3 H), 0.14 (d, 3 H, J ) 1.6 Hz); 13C
NMR (CDCl3) δ 171.7, 171.3, 163.74, 163.66, 158.8, 158.0,
J. Org. Chem, Vol. 70, No. 1, 2005 147