356 Journal of Medicinal Chemistry, 2006, Vol. 49, No. 1
Fletcher et al.
(400 MHz, CDCl3) δH 0.88 (6 H, t, J ) 6.8 Hz, 2 CH2CH3), 1.23-
1.66 (48 H, m, 24 CH2), 2.10-2.16 (8 H, m, 2 CH2CtCCH2),
2.26 (6 H, s, N(CH3)2), 2.28-2.32 (4 H, m, 2 CH2CO2), 2.44 (2 H,
34.5 (2 CH2CO2), 54.5 (N(CH3)3), 54.8 (CH3O), 63.6 (CH2N-
(CH3)2), 66.1 (CH2CHCH2N(CH3)3), 66.3 (CHCH2N(CH3)3), 80.4,
80.7 (2 CtC), 173.1, 173.5 (2 CO); m/z (FAB+) 659 ([M]+, 46),
658 ([M]+, 100); HRMS (FAB+) calcd for C42H76N1O4 [M]+
658.577436, found 658.579575; HPLC analysis method 1 tR ) 30.6
min (>98% pure), method 2 tR ) 21.95 min (>96% pure).
N-[1-(2,3-(Dioctadec-14-ynoyloxy))propyl]-N,N,N-trimethyl-
ammonium methyl sulfate (DS(14-yne)TAP, 4): 77% (205 mg);
Rf 0.37 (CH2Cl2:MeOH:H2O, 65:25:4); 1H NMR (400 MHz, CDCl3)
δH 0.90 (6 H, t, J ) 7.4 Hz, 2 CH2CH3), 1.15-1.34 (32 H, 2 m, 16
CH2), 1.36-1.48 (8 H, m, 4 CH2), 1.48-1.58 (4 H, m, 2 CH2CH2-
CO2), 2.02-2.10 (8 H, m, 2 CH2CtCCH2), 2.22-2.30 (4 H, m, 2
CH2CO2), 3.32 (9 H, s, N(CH3)3), 3.63 (3 H, s, CH3O), 3.66-3.71
2
3
m, C3-Ha,b), 4.08 (1 H, dd, J ) 11.6 Hz, J ) 6.4 Hz, C1-Hb),
2
3
4.36 (1 H, dd, J ) 11.8 Hz, J ) 3.0 Hz, C1-Ha), 5.19 (1 H, m,
C2-H); 13C NMR (400 MHz, CDCl3) δC 14.5 (2 CH2CH3), 19.1,
19.1, 23.0, 25.3, 29.0, 29.2, 29.3, 29.3, 29.4, 29.5, 29.5, 29.5, 29.6,
32.2 (26 CH2), 34.3, 34.5 (2 CH2CO2), 46.4 (N(CH3)2), 59.8 (CH2N-
(CH3)2), 64.2 (CH2CHCH2N(CH3)2), 69.6 (CHCH2N(CH3)2), 80.4,
80.7 (2 CtC), 173.5, 173.8 (2 CO); m/z (FAB+) 645 ([M + H]+,
30), 644 (M+, 70), 84 ([(CH3)2NCH2CtCH + H]+, 42), 58
([(CH3)2NdCH2]+, 100); HRMS (FAB+) calcd for C41H73N1O4
[M + H]+ 644.561786, found 644.561218.
2
3
(1 H, dd, J ) 14.0 Hz, J ) 8.8 Hz, C3-Ha/b), 3.99-4.04 (1 H,
1,2-Bis(octadec-14-ynoyloxy)-3-(dimethylamino)propane (DS-
(14-yne)DAP, 25): 424 mg (98%); Rf 0.37 (ethyl acetate); 1H NMR
(270 MHz, CDCl3) δH 0.90 (6 H, t, J ) 7.4 Hz, 2 CH2CH3), 1.17-
1.34 (34 H, 2 m, 17 CH2), 1.36-1.48 (6 H, m, 2 CH2CH3, CH2),
1.48-1.58 (4 H, m, CH2CH2CO2), 2.02-2.10 (8 H, m, 2 CH2Ct
CCH2), 2.19 (6 H, s, N(CH3)2), 2.21-2.26 (4 H, m, 2 CH2CO2),
2
3
dd, J ) 12.4 Hz, J ) 5.6 Hz, C1-Hb), 4.11-4.17 (1 H, br d,
2J ) 13.6 Hz, C3-Hb/a), 4.41-4.45 (1 H, dd, J ) 12.0 Hz, J )
3.6 Hz, C1-Ha), 5.49-5.59 (1 H, m, C2-H); 13C NMR (400 MHz,
CDCl3) δC 13.9 (2 CH2CH3), 19.1, 21.2, 22.9, 25.0, 25.1, 29.3,
29.5, 29.5, 29.6, 29.7, 29.7, 29.8, 29.9, 29.9, 30.0, 30.0, 30.0 (26
CH2), 34.2, 34.6 (2 CH2CO2), 54.5 (N(CH3)3), 54.8 (CH3O), 63.6
(CH2N(CH3)2), 66.2 (CH2CHCH2N(CH3)3), 66.3 (CHCH2N(CH3)3),
80.4, 80.8 (2 CtC), 173.6, 173.2 (2 CO); m/z (FAB+) 659 ([M +
H]+, 46), 658 ([M]+, 100); HRMS (FAB+) calcd for C42H76N1O4
[M]+ 658.577436, found 658.577713; HPLC analysis method 1
tR ) 30.7 min (>98% pure), method 2 tR ) 22.02 min (>99%
pure).
2
3
2
2.32-2.43 (2 H, m, C3-Ha,b), 3.99-4.04 (1 H, dd, J ) 11.9 Hz,
3J ) 6.4 Hz, C1-Hb), 4.27-4.32 (1 H, dd, 2J ) 11.9 Hz, 3J ) 3.1
Hz, C1-Ha), 5.09-5.17 (1 H, m, C2-H); 13C NMR (400 MHz,
CDCl3) δC 13.9 (2 CH2CH3), 19.1, 21.1, 22.9, 25.3, 25.3, 29.2,
29.5, 29.6, 29.7, 29.9, 29.9, 30.0, 30.0, (26 CH2), 34.5, 34.8, (2
CH2CO2), 46.4 (N(CH3)2), 59.8 (CH2N(CH3)2), 64.2 (CH2CHCH2N-
(CH3)2), 69.6 (CHCH2N(CH3)2), 80.4, 80.7 (2 CtC) 173.9, 173.6
(2 CO); m/z (FAB+) 645 ([M + H]+, 29), 644 (M+, 68), 84
([(CH3)2NCH2CtCH + H]+, 37), 58 ([(CH3)2NdCH2]+, 100);
HRMS (FAB+) calcd for C41H73N1O4 [M + H]+ 644.561786, found
644.56257.
Acknowledgment. Dr. Steven Fletcher gratefully acknowl-
edges funding from IC-Vec, Ltd. Funding for Dr. Ayesha
Ahmad provided by a National Science Foundation, USA, IRFP
grant. X-ray diffraction studies were carried out at both the
University of CaliforniasSanta Barbara (UCSB) facilities by
Dr. Cyrus Safinya and Dr. Heather Evans and at Imperial
College, London, by Dr. Richard Templer and Andrew Heron.
General Procedure for Preparation of Quarternary Amino
“TAP” Lipids as Their Methyl Sulfate Salts (2, 3, and 4). The
“DAP” lipids (23, 24, or 25; 1 equiv) were dissolved in anhydrous
acetone (0.1 M) and stirred under N2 at 0 °C. Then, a solution of
dimethyl sulfate (6 equiv) in anhydrous acetone (2 M) was added
dropwise. After stirring for 24 h at 4 °C, the reaction mixture was
filtered, washing with cold acetone to give the desired “TAP” lipids
in approximately 50% yields as white powders. Column chroma-
tography (CHCl3:MeOH:H2O, 85:13.4:1.6) of the filtrate typically
afforded around a further 25% yield of each “TAP” lipid.
N-[1-(2,3-(Dioctadec-4-ynoyloxy))propyl]-N,N,N-trimethylam-
monium methyl sulfate (DS(4-yne)TAP, 2): 75% (190 mg); Rf
0.39 (CH2Cl2:MeOH:H2O, 65:25:4); 1H NMR (400 MHz, CDCl3)
δH 0.81 (6 H, t, 3J ) 6.8 Hz, 2 CH2CH3), 1.15-1.30 (40 H, m, 20
CH2), 1.31-1.43 (4 H, m, 2 CH2(CH2)10CH3), 1.98-2.07 (4 H, m,
2 CH2(CH2)11CH3), 2.24 (6 H, s, N(CH3)2), 2.30-2.46 (4 H, m, 2
CH2CH2CO2), 2.46-2.55 (4 H, m, 2 CH2CH2CO2), 3.34 (9 H, s,
N(CH3)3), 3.63 (3 H, s, CH3O), 3.72-3.78 (1 H, dd, 2J ) 14.0 Hz,
3J ) 8.8 Hz, C3-Ha/b), 4.06-4.10 (1 H, dd, 2J ) 12.2 Hz, 3J ) 5.4
Hz, C1-Hb), 4.22-4.25 (1 H, br d, 2J ) 13.6 Hz, C3-Hb/a), 4.45-
Supporting Information Available: Table providing results
from purity analyses by HPLC, and 1H NMR and 13C NMR spectra
for key target compounds (2 - 4), and experimental and charac-
terization data for compounds 6-9 and 11-15. This material is
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2
3
4.49 (1 H, dd, J ) 12.0 Hz, J ) 3.6 Hz, C1-Ha), 5.51-5.59 (1
H, m, C2-H); 13C NMR (400 MHz, CDCl3) δC 14.5 (2 CH2CH3),
15.0, 15.0, 19.1, 19.1, 23.1, 29.3, 29.4, 29.4, 29.4, 29.6, 29.8, 30.0,
30.1, 30.1, 30.1, 32.3 (26 CH2) 34.1, 34.5 (2 CH2CO2), 54.6
(N(CH3)3), 54.8 (CH3O), 63.7 (CH2N(CH3)3), 66.1 (CH2CHCH2N-
(CH3)3), 66.7 (CHCH2N(CH3)3), 81.7, 82.2 (2 CtC), 171.7, 172.0
(2 CO); m/z (FAB+) 659 ([M]+, 46), 658 ([M]+, 100); HRMS
(FAB+) calcd for C42H76N1O4 [M]+ 658.577436, found 658.578583;
HPLC analysis method 1 tR ) 29.9 min (>98% pure), method 2
tR ) 20.82 min (>95% pure).
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N-[1-(2,3-(Dioctadec-9-ynoyloxy))propyl]-N,N,N-trimethylam-
monium methyl sulfate (DS(9-yne)TAP, 3): 200 mg (76%); Rf
0.37 (CH2Cl2:MeOH:H2O, 65:25:4); 1H NMR (400 MHz, CDCl3)
δH 0.81 (6 H, t, J ) 6.7 Hz, 2 CH2CH3), 1.15 - 1.59 (48 H, 2 m,
24 CH2), 2.04-2.13 (8 H, m, 2 CH2CtCCH2), 2.22-2.32 (4 H,
m, 2 CH2CO2), 3.29 (9 H, s, N(CH3)3), 3.63 (3 H, s, CH3O), 3.63-
3.72 (1 H, dd, 2J ) 14.4 Hz, 3J ) 8.8 Hz, C3-Ha/b), 3.98-4.09 (2
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2
3
H, m, C1-Hb, C3-Hb/a), 4.41-4.45 (1 H, dd, J ) 11.8 Hz, J )
3.0 Hz, C1-Ha), 5.52 (1 H, m, C2-H); 13C NMR (400 MHz, CDCl3)
δC 14.5 (2 CH2CH3), 19.1, 19.1, 23.0, 25.0, 25.1, 29.0, 29.1, 29.2,
29.2, 29.3, 29.4, 29.4, 29.5, 29.5, 29.5, 29.6, 32.2 (26 CH2), 34.2,