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ARVIND MISRA et al.
chemistry. The overall coupling yield was found more H, t, J 5.8 Hz, NHOCO), 3.28–3.34 (12 H, m, –NHCH2–
than 85% during the course of automated synthesis of oli- CH2–NH–), 2.85 (6 H, s, NCH3), 4.57 (1 H, br, SO2NH),
gonucleotides.
8.01 (1 H, t, naphthoyl), 8.52–7.87 (3 H, m, naphthoyl),
6.44–6.50 (2 H, m, H1', naphthoyl), 7.59 (1 H, d, J 7.9 Hz,
H6), 5.16 (1 H, d, J 8.1 Hz, H5), 5.68 (1 H, d, H1'), 5.49
(1 H, m, H2'), 4.87 (1 H, m, H3'), 3.91 (1 H, m, H4'), 3.60
(2 H, m, H5'), 0.87–1.24 (28 H, m, Pri).
3',5'-O-(tetraisopropyldisiloxane-1,3-diyl)uridine (1).
3',5'-OH of uridine was first protected by treating with sily-
lating reagent 1,1,3,3-tetraisopropyldisiloxane-1,3-dichlo-
ride in pyridine under anhydrous condition to give (1) (yield
75%) as described previously [5].
2'-O-(Dansyl-6-sulphonamidohexylaminocarbo-
nyl)uridine (5). Compound (4) (3 mM, 2.58 g) was dis-
solved in THF (10 ml) and 1 M solution of TBAF in THF
(5.0 ml) was added with stirring for 20 min. The reaction
mixture was poured in 5% NaHCO3 solution and extract-
ed several times with ethyl acetate. The organic layer was
concentrated, dried, filtered and again evaporated to af-
ford the compound as a gummy mass. Yield 1.76 g (95%).
Rf 0.37 (CHCl3/MeOH, 7 : 3), MALDI-TOF (2.5-DH-
BA): [M + H]+ calcd. 619.4, found 618.6, [M + Na]+ calcd.
642.4, found 643.3, [M + K]+ calcd. 658.4, found 659.3.
1H NMR (δ, ppm): 11.38 (1 H, s, H3), 7.11 (1 H, t, J 5.8
Hz, NHOCO), 3.28–3.34 (12 H, m, –NHCH2–CH2–NH–),
2.85 (6 H, s, NCH3), 4.57 (1 H, br, SO2NH), 8.01 (1 H, t,
naphthoyl), 8.52–7.87 (3 H, m, naphthoyl), 6.44–6.50
(2 H, m, H1', naphthoyl), 7.59 (1 H, d, J 7.9 Hz, H6), 5.16
(1 H, d, J 8.1 Hz, H5), 5.68 (1 H, d, H1'), 5.49 (1 H, m,
H2'), 4.87 (1 H, m, H3'), 3.91 (1 H, m, H4'), 5.57 (1 H, m,
3'-OH), 5.21 (1 H, m, 5'-OH), 3.60 (2 H, m, H5').
3'-O-(N,N-Diisopropylamino-2-cyanoethoxyphos-
phinyl)-5'-O-(4,4'-dimethoxytrityl)uridine-2'-yl carba-
mates (7). Uridin-2'-yl carbamate was first treated with
4,4'-dimethoxytrityl chloride (DMTr-Cl) in presence of
dimethylaminopyrimidine (DMAP) and TEA to get 5'-O-
(4,4'-dimethoxytrityl) uridin-2'-yl carbamate (6). To the
well-stirred solution of (6) (1.3 mM, 1.2 g) in anhydrous
dichloromethane (12 ml), 2-cyanoethyl-N,N,N',N''-tetrai-
sopropylphosphoramidite (Bis-reagent) (3.5 mM, 1.12 ml)
was added at ambient temperature. Pyridinium trifluoroac-
etate (Py · TFA) (3.5 mM, 0.82 g) was added to the reaction
mixture and stirred for 3 h. After complete consumption of
starting material (monitored on TLC), methanol (3 ml) was
added, and the resulting solution was washed with 10%
NaHCO3 (3 × 15 ml). Organic layer was dried over sodium
sulfate and directly applied to a short silica gel column.
Compound was eluted with dichloromethane/ethyl ace-
2'-O-(Imidazol-1-ylcarbonyl)-3',5'-O-(tetraisopro-
pyldisiloxane-1,3-diyl)uridine (2). The compound (1)
(5 mM, 2.434 g) was evaporated twice with anhydrous ac-
etonitrile (2 × 20 ml) and dissolved in dry dicloromethane
(25 ml). N,N'-carbonyldiimidazole (CDI) (6.0 mM, 1 g)
was added. The reaction mixture was stirred for 2 h. TLC
showed the conversion of starting material into product of
lower mobility. The solution was washed with water
(2 × 30 ml), dried over sodium sulphate, filtered and evap-
orated in vacuum to get imidazolyl derivative as white
foam. Yield 2.24 g (91.9%). Rf 0.41 (EtOAc). MALDI-
TOF (2,4,6-THAP + citrate): [M + Na]+ calcd. 603.77,
found 604.58, [M + K]+ calcd. 619.88, found 620.55.
1H NMR (δ, ppm): 11.3 (1 H, s, H3), 8.3 (1 H, s, imida-
zole), 7.69 (1 H, s, imidazole), 7.93 (1 H, d, J 7.8 Hz,
H6), 6.97 (1 H, s, imidazole), 5.75 (1 H, s, H1'), 5.67
(1 H, d, J 5.2 Hz, H2'), 5.65 (1 H, d, J 8.0 Hz, H5), 4.71
(1 H, m, H3'), 4.23–3.95 (3 H, m, H4', H5'), 1.12–0.9
(28 H, m, Pri).
6-Aminohexyldansylsulphonamide (3). Dansyl chlo-
ride (5.0 mM, 1.35 g) was taken in THF (25 ml) and added
dropwise to a well stirred ice cold 10% NaHCO3 aqueous
solution of hexamethylene-1,6-diamine (20.0 mM) for 3 h.
Completion of reaction was monitored on TLC. The crude
product was poured in water and extracted with ethyl ace-
tate (3 × 25 ml). The organic layer was pooled, concentrat-
ed in vacuum and eluted with EtOAc/hexane (2 : 8) on sil-
ica gel column to get 6-aminohexyldansylsulphonamide
(3). Yield 2.33 g (78%). Rf 0.34 (8 : 2 : DCM/MeOH). UV
λ
max (MeOH) : 251, 334, 385 nm. 1H NMR (δ, ppm); 2.38–
2.69 (12 H, m, (–CH2)6NH2), 2.89 (6 H, s, (CH3)2N), 4.62
(3 H, br, NH), 8.21 (1 H, d, J 8.1 Hz, H2) 7.67–8.42 (3 H,
m, naphthoyl) 6.44–6.50 (2 H, m, naphthoyl). λmax 335 nm,
λem 535 nm (in methanol).
2'-O-(Dansyl-6-sulphonamidohexylaminocarbon- tate/Et3N (4.5 : 4.5 : 1, v/v/v). The desired fractions were
yl)-3',5'-O-(tetraisopropyldisiloxane-1,3-diyl)uridine (4). pooled and concentrated under reduced pressure to afford
Imidazolyl derivative of uridine (3.5 mM, 2 g) was taken title compound as amorphous yellow powder. Yield 1.24 g
in dry dichloromethane (30 ml) and (3) (4.5 mM, 1.58 g) (85%). Rf 0.63 (CH2Cl2/EtOAc/Et3A: 4.5 : 4.5 : 1, v/v/v).
was added dropwise for 1 h under argon atmosphere. Af-
λ
max 256, 335 nm. 1H NMR (δ, ppm): 11.42 (1 H, s, NH),
ter complete addition the reaction mixture was allowed to 8.39 (1 H, d, J 9.1 Hz, naphthoyl), 8.22–8.27 (2 H, m,
stir overnight. The reaction mixture was washed with wa- naphthoyl + H6), 7.59–7.96 (2 H, d, J 9 Hz, naphthoyl),
ter and 3% citric acid. The organic layer was kept over so- 7.28–7.37 (7 H, m, naphthoyl, C5, Ar), 7.08–7.78 (8 H, d,
dium sulphate and evaporated in vacuum. The residue was J 8.5 Hz, DMTr), 7.53 (1 H, t, J 5.9 Hz, NHCOO), 6.65
applied to silica column and product was isolated using (1 H, d, J 9 Hz, naphthoyl), 6.39–6.43 (1 H, m, H1'), 5.03
gradient ethyl acetate in chloroform. Fractions containing (1 H, m, H2'), 4.57 (1 H, br, SO2NH), 4.14–4.17 (2 H, m,
the product were evaporated in vacuum to give compound H3', H4'), 4.01–4.03 (2 H, m, H5'), 3.91 (2 H, t, P–O–CH2),
(4) as white solid foam. Yield 2.82 g (94%). Rf 0.45 3.87 (6 H, s, –OCH3), 3.21–3.32 (4 H, m, NHCH2), 3.14
(DCM/MeOH: 8 : 2 v/v). MALDI-TOF (2,5-DHBA): [M (2 H, m, NCH-), 2.93 (6 H, s, –NCH3), 2.62 (2 H, t,
+ Na]+ calcd. 884.3, found 885.2, [M + K]+ calcd. 900.3, CH2CN), 1.92–1.71 (8 H, m, NHCH2(CH2)4CH2NH),
found 900.9. 1H NMR (δ, ppm): 11.38 (1 H, s, H3), 7.11 (1 1.22 (12 H, d, -NCH(CH3)2). MALDI-TOF (2.5-DHAP):
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 35 No. 1 2009