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A.E. Beilstein, M.W. Grinstaff / Journal of Organometallic Chemistry 637–639 (2001) 398–406
2.1.5. Ferrocenoyl butanolamide phosphoramidite (8)
Following a published procedure from a German
Patent [49], ferrocenoylbutanolamide (0.100 g, 0.33
mmol) was azeotroped from dry Py. The solid was then
dissolved in 5 ml THF and 0.23 ml (1.32 mmol) of
DIEA was added. Next, 0.12 ml (0.50 mmol) of b-cy-
anoethylchloro-N,N-diisopropylphosphoramidite was
added and the mixture was stirred for 15 min. Water
(33 ml) was added to the flask and the mixture was
stirred for an additional 30 min. Ether–TEA was added
to the mixture, causing a yellow precipitate to form and
then 15 ml of 10% sodium bicarbonate was added,
dissolving the precipitate. The mixture was extracted
and then washed twice with water. The organic layer
was dried over sodium sulfate, then filtered and dried
by rotary evaporation. 31P-NMR (CDCl3) indicates a
peak at 148.46 ppm.
oration to yield 0.222 g solid (40% yield). 1H-NMR
(Me2SO, l ppm): 1.78 (s, 3H, 5-methyl); 2.08–2.18 (m,
2H, C2%); 2.75 (s, 2H, amine); 3.37 (s, 2H, C5%); 3.75 (q,
1H, C4%); 4.20 (m, 1H, C3%); 5.20 (s, 1H, 3%-OH); 6.20 (t,
1H, C1%); 7.68 (s, 1H, C6). FAB-MS: m/z: [M+H] 242.
2.1.9. N-Ferrocenoyl-5%-amido-5%-deoxythymidine (13)
A solution of 0.220 g (0.913 mmol) of 12, 0.230 g (1
mmol) of 1, 0.135 g of HOBt (1 mmol), and 0.210 g of
DCC (1.03 mmol) in 25 ml dry DMF was prepared.
TEA (0.14 ml) was added to the solution. After 12 h,
DCM was added and the mixture was filtered to re-
move a yellow solid from the red–orange filtrate. The
yellow powder was dissolved in 50 ml THF and chilled
in an ice bath for 2 h, then filtered to remove urea. The
resulting filtrate was dried to yield a red solid, which
was purified by column chromatography (40% yield).
1H-NMR (ACN, l ppm)1.85 (s, 3H, Me); 2.20 (t, 2H,
2% protons); 3.45–3.62 (m, 1H, 5% protons); 3.94 (m, 1H,
4% proton); 4.16 (s, 5H, Cp ring, unsubstituted); 4.29 (m,
1H, 3% proton); 4.35 (t, 2H, Cp, m); 4.72 (t, 2H, Cp, o);
6.13 (t, 1H, 1% proton); 6.86 (bs, 1H, NH); 7.41 (s, 1H,
C6H); 9.30 (s, 1H, N3H). ESI MS: Found, 453.22.
Calculated for C21H23FeN3O5, 453.3.
2.1.6. 5%-O-Methanesulfonyl-thymidine (10)
The reaction was based on a literature procedure
[50]. Thymidine 9 (1.53 g, 6.3 mmol) was dissolved in
10 ml of dry Py and cooled to −10 °C. Next,
methanesulfonyl chloride (0.5 ml dissolved in 3 ml
DCM) was added dropwise over a period of 20 min.
The reaction mixture was then kept at 0 °C. After 16 h,
10 ml of MeOH was added to quench the reaction and
the solvents were evaporated via high vacuum. The
resulting crude product was checked by TLC and
purified by column chromatography (silica gel,
CH3OH–CHCl3 (1:19)). A white, powdered solid 10
2.1.10. N-ferrocenoyl-5%-amido-5%-deoxythymidine-3%-
phosphoramidite (14)
A solution of 0.095 g (0.2 mmol) of 13 was
azeotroped from Py twice. DIEA (0.14 ml, 0.8 mmol)
and 3 ml of THF were added to the flask. Once
dissolved, 70 ml (0.31 mmol) of b-cyanoethylchloro-
N,N-diisopropylphosphoramidite was added. The mix-
ture was stirred until the solution lost turbidity (ꢀ15
min). The solvent was removed in vacuo. 31P-NMR
(CD3CN) indicated chemical shifts at 150.6 and 149.5
ppm, corresponding to the nucleoside phosphoramidite.
1
was obtained (1.101 g; 55% yield). H-NMR (Me2SO, l
ppm): 1.78 (s, 3H, 5-methyl); 2.08–2.22 (m, 2H, C2%);
3.22 (s, 3H, CH3S); 3.98 (q, 1H, C4%); 4.28 (m, 1H, C3%);
4.40 (m, 2H, C5%); 5.50 (s, 1H, 3%-OH); 6.22 (t, 1H, C1%);
7.48 (s, 1H, C6); 11.25 (s, 1H, N3). FAB-MS: m/z:
[M+H]+ 321.
2.1.7. 5%-Azido-5%-deoxythymidine (11)
2.2. Oligodeoxynucleotide syntheses
A solution of 10 (1.00 g, 3.125 mmol) in 20 ml of
DMF containing lithium azide (0.555 g, 11.3 mmol)
was stirred at 90 °C under nitrogen. After 3 h, the
solvent was removed under vacuum. The resulting
crude product was purified by column chromatography
(silica gel, CH3OH–CHCl3 (1:19)) to afford 11 (0.626 g;
75% yield). 1H-NMR (Me2SO, l ppm): 1.78 (s, 3H,
5-methyl); 2.08–2.22 (m, 2H, C2%); 3.57 (d, 2H, C5%);
3.85 (q, 1H, C4%); 4.20 (m, 1H, C3%); 5.50 (s, 1H,
3%-OH); 6.22 (t, 1H, C1%); 7.48 (s, 1H, C6); 11.25 (s, 1H,
N3). FAB-MS: m/z: [M+H]+ 268.
2.2.1. On-column deri6atization
Syntheses were conducted on the 1.0 mM scale. Stan-
dard automated DNA synthesis was performed from
the 3%- to 5%-terminus until a halogenated nucleoside
(8-bromoadenosine or 5-iodouridine) was incorporated.
The synthesis was interrupted prior to cleavage of the
5%-dimethoxytrityl group, leaving the ODN bound to
the column. N2 was blown through the column for 15
min to dry the column. Next, the column was uncapped
and 20 mg FPA, 10 mg Pd(PPh3)4, and 1 mg CuI were
added with 30% TEA in DMF. With closed stopcocks
on each end of the column, the column was sealed and
was shaken on a hand-shaker for 6 h. The column was
then rinsed by pushing through 10 ml of the TEA–
DMF mixture, then 50 ml of MeCN. The column was
dried for 15 min under N2. Finally, the column was
placed back on the synthesizer and standard automated
2.1.8. 5%-Amino-5%-deoxythymidine (12)
To a solution of 5%-azidothymidine (11), (0.6258 g,
2.2 mmol) in 30 ml MeOH was added an excess of 10%
Pd/C. H2 (g) was bubbled through the mixture at 1 atm
for 4 h. The mixture was then filtered through Celite to
remove Pd/C and the filtrate was dried by rotary evap-