A R T I C L E S
Schwo¨rer and Schmidt
3
3
butyl hydroperoxide (270 µL, 1.49 mmol, 5.5 M in nonane) was added.
After 2 h, triethylamine (5 mL) was added, and the solution was stirred
overnight. Evaporation of the solvents and purification by flash
chromatography (ethyl acetate/methanol 5:1 v/v + 1% triethylamine)
afforded the triethylammonium salts (R,S)-14a (600 mg, 82%, mixture
of diastereomers) as a colorless lyophilisate. Rf ) 0.17 (ethyl acetate/
methanol 5:1 v/v + 1% triethylamine); 1H NMR (250 MHz, D2O): δ
5.80 (d, J(1′,2′) ) 3 Hz, 1 H, 1′-H), 6.03 ((d, J(5,6) ) 7.8 Hz, 1 H,
3
5-H), 7.01-7.28 (m, 5 H, Ph), 8.09 (d, J(5,6) ) 7.8 Hz, 1 H, 6-H);
13C NMR (150.9 MHz, D2O): δ ) 52.2 (2′′-C), 63.5 (2J(C,P) ) 4.6
Hz; 5′-C), 65.3 (3′′-C), 68.4 (3′-C), 74.6 (2′-C), 73.4 (1J(C,P) ) 160
Hz, 6′′-C), 82.6 (4′-C), 89.7 (1′-C), 95.1 (6-C), 98.4 (1′′-C), 103.7 (4′′-
C), 116.8/123.2/129.8 (Ph), 143.9 (5-C), 148.2 (5′′-C), 156.7 (4-C),
174.5 (2-C); 31P NMR (243 MHz, D2O): δ ) 1.52 (bd, J(P,P) ) 25
3
3
3
) 1.07 (2t, J ) 7.3 Hz, 9 H; 3 ethyl), 1.81/1.86/1.94/1.97 (8s, 12 H;
Hz, PO4), 13.44 (bd, J(P,P) ) 25 Hz, PO3); MALDI-MS (negative
NHAc, 3 acetyl), 3.00 (2q, 3J ) 7.3 Hz, 6 H; 3 ethyl), 3.75-4.07 (m,
3 H; 4′-H,5′a-H, 5′b-H), 4.25-4.51 (m, 6 H; 2′′-H, 3′′-H, allyl), 4.77
mode, matrix: ATT): m/z (%): 663 (100) [M - 3Na+ + 2H+]-, 730.4
for C23H27N4Na3O15P2.
1al. HPLC: prep. RP-18, column A (flow 15 mL min-1, 0.05 M
TEAB buffer, 6% acetonitrile): tR ) 11.5 min; H NMR (600 MHz,
D2O): δ ) 1.94 (s, 3H; NHAc), 4.01 (m, J(5′a,5′b) ) 11.4 Hz, 1 H;
2
3
(dd, J(6′′,P) ) 11.7 Hz, J(6′′,P) ) 15.9 Hz, 1 H; 6′′-H), 5.02-5.27
(m, 6 H; 2′-H, 3′-H, allyl), 5.39-5.48 (m, 2 H; 1′′-H, 4′′-H), 5.61 (d,
3J(1′,2′) ) 5.3 Hz, 1 H; 1′-H), 5.66-5.84 (m, 2 H; allyl), 6.75-7.51
(m, 8 H; Ph, Bz), 6,91 (d, 3J(5,6) ) 7.6 Hz, 1 H; 5-H), 7.75-7.84 (m,
2 H; Bz), 7.96 (d, 3J(5,6) ) 7.6 Hz, 1 H; 6-H); MALDI-MS (negative
mode, matrix: ATT): m/z (%): 973 (15) [M - HNEt3]-, 933 (100)
1
2
5′a-H), 4.16 (m, 1 H; 2′-H), 4.19 (m, 1 H; 4′-H), 4.22 (m, 1 H; 3′-H),
4.28 (m, 2J(5′a,5′b) ) 11.4 Hz, 1 H; 5′b-H), 4.33 (m, 1 H; 3′′-H), 4.37
(m, 3J(1′′,2′′) ) 7.3 Hz, 1 H; 2′′-H), 4.41 (m, 2J(6′′,P) ) 12.9 Hz, 1 H;
6′′-H), 5.20 (dd, 3J(3′′,4′′) ) 4J(4′′,6′′) ) 3.2 Hz, 1 H; 4′′-H), 5.43 (d,
3J(1′′,2′′) ) 7.3 Hz, 1 H, 1′′-H), 5.88 (d, 3J(1,2) ) 2.5 Hz, 1 H, 1′-H),
6.00 (d, 3J(5,6) ) 7.9 Hz, 1 H, 5-H), 6.99-7.26 (m, 5 H, Ph), 8.12 (d,
3J(5,6) ) 7.9 Hz, 1 H, 6-H); 13C NMR (150.9 MHz, D2O): δ ) 52.5
(2′′-C), 63.4 (5′-C), 66.26 (3′′-C), 67.9 (3′-C), 71.6 (1J(C,P) ) 160
Hz, 6′′-C), 74.6 (2′-C), 82.1 (4′-C), 89.9 (1′-C), 94.7 (6-C), 98.4 (1′′-
C), 101.5 (4′′-C), 116.5/123.2/129.8 (Ph), 144.2 (5-C), 149.4 (5′′-C),
158.4 (4-C), 174.6 (2-C); 31P NMR (243 MHz, D2O): δ ) 1.45 (bd,
+
[M - HNEt3 - All+ + H+]-; MALDI-MS (positive mode, matrix:
+
DHB): m/z (%): 997 (100) [M - HNEt3 + Na+ + H+]+, 875 (80)
[M - HNEt3+ - PhCOOH + Na+ + H+]+, 1075.4 for C48H63N4O19P2.
Triethylammonium [2-Benzoyloxyethyl (6R,S)-2-acetamido-3-
O-benzoyl-2,4-dideoxy-6-diallylphosphoryl-â-L-threo-hex-4-eno-
pyranoside-6-yl](N-acetyl-2′,3′-di-O-acetylcytidine-5′-yl)phosphate
(14b). Alcohol 12b (140 mg, 0.23 mmol) and cytidine phosphoramidite
1319(160 mg, 0.28 mmol) were coevaporated with dry dichloromethane
and dried under high vacuum. The mixture was dissolved in dry
dichloromethane (2 mL) and tetrazole (32 mg, 0.44 mmol) was added.
After stirring for 3 h at room temperature, the coupling reaction was
complete, and tert-butyl hydroperoxide (70 µl, 0.38 mmol, 5.5 M in
nonane) was added. After 2 h, triethylamine (4 mL) and dichloro-
methane (2 mL) were added and the solution stirred overnight.
Evaporation of the solvents and purification by flash chromatography
(ethyl acetate/methanol 5:1 v/v + 1% triethylamine) afforded the
triethylammonium salts 14b (140 mg, 55%, mixture of diastereomers)
as a colorless lyophilisate. Rf ) 0.78 (ethyl acetate/methanol 1:1 v/v
3J(P,P) ) 25 Hz, PO4), 13.34 (bd, J(P,P) ) 25 Hz, PO3); MALDI-
3
MS (negative mode, matrix: ATT): m/z (%): 663 (100) [M - 3Na +
2H]-; FAB-MS (positive mode, matrix: glycerol/acetonitrile/0.1%
trifluoroacetic acid 1:1:1): 687 (20) [M - 2Na+ + 3H+]+, 709 (40)
[M - Na+ + 2H+]+, 725 (35) [M - 2Na+ + K+ + 2H+]+, 731 (35)
[M + H+]+, 730.4 for C23H27N4Na3O15P2.
Trisodium [2′-Hydroxyethyl (6R,S)-2-acetamido-3-O-benzoyl-
2,4-dideoxy-6-phosphoryl-â-L-threo-hex-4-enopyranoside-6-yl]-
(cytidine-5′-yl)phosphate (1bh,l). A solution of 14b (30 mg, 0.026
mmol) in aqueous ammonia (25%, 4 mL) was stirred for 3 days at
room temperature. The mixture was concentrated and lyophilized from
water. To a solution of the resulting lyophilisate and dimedone (50
mg, 0.37 mmol) in THF (2.5 mL), was added Pd(PPh3)4 (15 mg, 12
µmol) in the dark. After 5 h the solvents were evaporated and dimedone
was removed by RP-18 chromatography (ethanol/water 1:3 v/v). After
lyophilization from water, separation of diastereomers by RP-18 HPLC,
conversion into the sodium salts by IR 120 (Na+) and lyophilization
from water 1bh (2 mg, 11%) and 1bl (5 mg, 27%) were collected.
1
+ 1% triethylamine); H NMR (250 MHz, D2O): δ ) 1.30 (2t, 9 H;
3 N-ethyl), 1.92-2.15 (8s, 12 H; NHAc, 3 acetyl), 3.00 (2q, 6 H; 3
N-ethyl), 3.85-4.7 (m, 11 H; 1′′′a-H, 1′′′b-H, 2′′′a-H, 2′′′b-H, 4′-H,
5′a-H, 5′b-H, allyl), 4.9-5.7 (m, 11 H; 1′-H, 2′-H, 3′-H, 2′′-H, 3′′-H,
3
4′′-H, 6′′-H, allyl), 5.84-6.00 (m, 2 H; allyl), 6.17 (2 d, J(1′′,2′′) )
4.3 Hz, 1 H; 1′′-H), 7.15-7.58 (m, 6 H; 5-H, Bz), 7.90-8.00 (m, 4 H;
Bz), 8.40/8.44 (2d, 3J(5,6) ) 7.6 Hz, 1 H; 6-H); MALDI-MS (negative
+
mode, matrix: ATT): m/z (%): 1045 (40) [M - HNEt3
]
- 1004 (100)
+
[M - HNEt3 - All+ + H+]-; MALDI-MS (positive mode, matrix:
DHB): m/z (%): 1069 (50) [M - HNEt3+ + Na+ + H+]+, 969 (100)
1bh. HPLC: prep. RP-18 column (flow 15 mL min-1, 0.05 m TEAB
buffer, 1% acetonitrile): tR ) 11 min; H NMR (250 MHz, D2O): δ
1
[M - HNEt3 - PhCOOH + 2Na+]+, 1147.4 for C51H67N4O21P2.
+
) 1.92 (s, 3H; NHAc), 3.45-4.30 (m, 12 H; 1′′′a,b-H,2′′′a,b-H, 3′′-
H, 2′′-H, 5′a,b-H, 4′-H, 2′-H, 3′-H, 6′′-H), 4.95 (d, 1 H; 1′′-H), 5.20
(m, 1 H; 4′′-H), 5.84 (d, 3J(1′,2′) ) 3.5 Hz, 1 H; 1′-H), 6.16 (d, 3J(5,6)
Trisodium [Phenyl (6R,S)-2-acetamido-3-O-benzoyl-2,4-dideoxy-
6-phosphoryl-â-L-threo-hex-4-enopyranoside-6-yl](cytidine-5′-yl)-
phosphate (1ah,l). A solution of 14a (50 mg, 0.046 mmol) in aqueous
ammonia (25%, 3 mL) was stirred for 4 h at room temperature. The
mixture was concentrated and lyophilized from water. To a solution
of the resulting lyophilisate and dimedone (30 mg, 0.22 mmol) in THF
(2 mL) was added Pd(PPh3)4 (15 mg, 12 µmol) in the dark. After 5 h
the solvents were evaporated and dimedone was removed by RP-18
chromatography (ethanol/water 1:3 v/v). After lyophilization from water,
purification and separation of diastereomers by RP-18 HPLC (0.05 M
TEAB) the products were converted into their sodium salts by IR 120
(Na+) and lyophilized to yield 1ah (12 mg, 36%) and 1al (3 mg, 9%).
3
) 7.8 Hz, 1 H; 5-H), 8.02 (d, J(5,6) ) 7.8 Hz, 1 H; 6-H); MALDI-
MS (negative mode, matrix: ATT): m/z (%): 631 (100) [M - 3Na+
+ 2H+]-, 698.4 for C19H27N4Na3O16P2.
1bl. HPLC: prep. RP-18 column (flow 15 mL min-1, 0.05 M TEAB
1
buffer, 1% acetonitrile): tR ) 12 min; H NMR (600 MHz, D2O): δ
) 1.94 (s, 3H; NHAc), 3.64-3.90 (m, 4 H; 1′′′a,b-H,2′′′a,b-H), 4.02-
4.04 (m, 2 H; 2′′-H, 3′′-H), 4.10-4.20 (m, 2J(5′a,5′b) ) 12.7 Hz, 3 H;
5′a,b-H, 4′-H), 4.26-4.27 (m, 2 H; 2′-H, 3′-H), 4.45 (dd, 1J(6′′,P) )
4
3
12 Hz, J(4′′,6′′) ≈ 5 Hz), 1 H; 6′′-H), 5.01 (d, J(1′′,2′′) ) 5.6 Hz, 1
3
4
1ah. HPLC: prep. RP-18, column A (flow 15 mL min-1, 0.05 M
H; 1′′-H), 5.12 (dd, J(3′′,4′′) ≈ J(4′′,6′′) ≈ 5 Hz, 1 H; 4′′-H), 5.85
(d, 3J(1′,2′) ) 3.6 Hz, 1 H; 1′-H), 6.23 (d, 3J(5,6) ) 7.8 Hz, 1 H; 5-H),
8.11 (d, 3J(5,6) ) 7.8 Hz, 1 H; 6-H); 13C NMR (150.9 MHz, D2O): δ
) 52.4 (2′′-C), 60.5 (2′′′-C), 63.9 (2J(C,P) ) 4.6 Hz; 5′-C), 65.3 (3′′-
C), 68.7/73.4 (2′-C,3′-C), 70.7 (1′′′-C), 72.6 (1J(C,P) ) 155 Hz, 2J(C,P)
) 9.2 Hz; 6′′-C), 83.1 (4J(C,P) ) 7.8 Hz; 4′-C), 89.6 (1′-C), 95.4 (6-
C), 99.2 (1′′-C), 101.3 (3J(C,P) ) 7.5 Hz; 4′′-C), 143.5 (5-C), 149.3
(2J(C,P) ) 72 Hz; 5′′-C), 160.0 (4-C), 174 (2-C); 31P NMR (243 MHz,
D2O): δ ) 1.23 (b, PO4), 12.9 (b, PO3); MALDI-MS (negative mode,
1
TEAB buffer, 6% acetonitrile): tR ) 10 min; H NMR (600 MHz,
2
D2O): δ ) 1.95 (s, 3H; NHAc), 3.93 (m, J(5′a,5′b) ) 14 Hz, 1 H;
5′a-H), 4.09 (dd, 3J(1′,2′) ) 3 Hz, 3J(2′,3′) ) 4.8 Hz, 1 H; 2′-H), 4.12
(m, 3J(3′,4′) ) 4.8 Hz, 1 H; 4′-H), 4.14 (m, 2J(5′a,5′b) ) 14 Hz, 1 H;
3
3
5′b-H), 4.19 (dd, J(2′,3′) ) 4.8 Hz, J(3′,4′) ) 4.8 Hz, 1 H; 3′-H),
3
3
4.22 (m, J(2′′,3′′) ) 6.1 Hz, 1 H; 3′′-H), 4.36 (dd, J(2′′,3′′) ) 6.1
Hz, 3J(1′′,2′′) ) 6.4 Hz, 1 H; 2′′-H), 4.61 (m, 2J(6′′,P) ) 12.1 Hz, 1 H;
3
6′′-H), 5.19 (m, 1 H; 4′′-H), 5.54 (d, J(1′′,2′′) ) 6.4 Hz, 1 H, 1′′-H),
9
1636 J. AM. CHEM. SOC. VOL. 124, NO. 8, 2002