312
T. Tichý et al. / European Journal of Medicinal Chemistry 55 (2012) 307e314
4.3.2. 3-(Hexadecyloxypropyl) ester of 9-[2-(phosphonomethoxy)
ethyl]guanine (6)
ESI-HRMS calcd for C47H90N4O8P 869.6491, found: 869.6490
[M þ H]þ; calcd for C47H89N4O8PNa 891.6310, found: 891.6311
Yield: 63 mg (54%). Mp 243e247 ꢁC. Anal. Calcd for
C28H53N5O6.5P (hemihydrate): C, 56.55; H, 8.98; N, 11.78; P, 5.21.
Found: C, 56.92; H, 9.31; N, 11.42; P, 4.95. ESI-MS, m/z: 586.4 (97)
[M þ H]þ, 608.4 (100) [M þ Na]þ, 630.3 (51), 652.3 (32). ESI-HRMS
calcd for C28H53N5O6P 586.3728, found: 586.3726 [M þ H]þ; calcd
for C28H52N5O6PNa 608.3547, found: 608.3542 [M þ Na]þ. 1H NMR
[M þ Na]þ. 1H NMR (CDCl3 þ MeOD, ppm)
d
:0 0.89 (t, 6H,
0
0
0
0
J23 e22 ¼ 7.0, H-23 ),1.23e1.35 (m, 52H, H-10 to H-22 ), 1.56 (m, 4H,
H-90), 1.92 (m, 4H, H-60), 3.42 (m, 4H, H-70), 3.50 (m, 4H, H-80), 3.61
(dd, 1H, Jgem ¼ 12.3, J3 e2 ¼ 4.7, H-30b), 3.69 (dd, 1H, Jgem ¼ 12.3,
0
0
J3 e2 ¼ 4.8, H-30a), 3.87 (m, 1H, H-20), 3.89 (dd, 1H, Jgem ¼ 13.8,
0
0
J4 eP ¼ 9.1, H-40b), 4.01 (dd, 1H, Jgem ¼ 13.8, J4 eP ¼ 8.9, H-40a),
0
0
(D2O þ NaOD, ppm)
d
: 0.87 (t, 3H, J23 e22 ¼ 7.0, H-230), 1.22e1.31
4.10e4.18 (m, 4H, H-50), 4.37 (dd, 1H0, Jgem ¼ 14.6, J1 e2 ¼ 7.0, H-1 b),
0
0
0
0
0
(m, 26H, H-100 to H-220), 1.54 (m, 2H, H-90), 1.84e1.91 (m, 4H, H-
4.49 (dd, Jgem ¼ 14.6, J1 e2 ¼ 4.0, H-1 a), 8.00 (s, 1H, H-2), 8.04 (s,1H,
0
0
40, H-60), 3.39 (t, 2H, J8 e9 ¼ 7.0, H-80), 3.50 (t, 2H, J7 e6 ¼ 6.6, H-7 ),
H-8). 13C NMR (CDCl3 þ MeOD, ppm)
d
: 12.97 (C-230), 21.91 (C-220),
0
0
0
0
0
3.64 (m, 2H, H-30), 3.72 (br t, 2H, J2 e1 ¼ 5.1, H-20), 3.87 (q, 2H,
25.43 (C-100), 28.63e28.96 (m, C-90, C-110 to C-200), 29.97 (d,
0
0
0
J5 e6 ¼ J5 eP ¼ 6.5, H-50), 4.10 (br t, 2H, J1 e2 ¼ 5.1, H-1 ), 7.64 (s, 1H,
J6 eP ¼ 6.0, C-60), 29.98 (d, J6 eP ¼ 6.0, C-60), 31.21 (C-210), 43.65 (C-
0
0
0
0
0
0
0
H-8). 13C NMR (D2O þ NaOD, ppm)
d
: 14.59 (C-230), 23.22 (C-220),
10), 59.64 (C-30), 62.61 (d, J4 eP ¼ 167.6, C-40), 63.39 (d, J5 eP ¼ 6.7, C-
0
0
26.68 (C-100), 27.80 (d, J4 eP ¼ 132.1, C-40), 30.01, 30.17, 30.29, 30.49,
50), 63.55 (d, J5 eP ¼ 6.6, C-50), 65.66, 65.68 (C-70), 70.48 (C-80), 80.16
0
0
30.54, 30.58, 30.60, 30.63 (C-90, C-110 to C-200), 31.15 (d, J6 eP ¼ 6.3,
(d, J2 eP ¼ 11.3, C-20), 123.14 (C-5), 140.79 (C-8), 144.61 (C-2), 148.38
0
0
C-60), 32.65 (C-210), 43.38 (C-10), 61.91 (d, J5 eP ¼ 5.5, C-50), 66.79 (C-
(C-4), 156.85 (C-6).
0
30), 67.98 (C-70), 68.79 (C-20), 71.56 (C-80), 117.83 (C-5), 138.91 (C-8),
151.95 (C-4), 160.88 (C-2), 167.82 (C-6).
4.5.2. Bis 3-(hexadecyloxypropyl) ester of 9-(S)-[3-hydroxy-2-
(phosphonomethoxy)propyl]guanine (14)
4.4. General method D
Yield 0.68 g (32%). Mp 166e168 ꢁC. Anal. Calcd for C47H92N5O9P
(monohydrate): C, 62.57; H, 10.28; N, 7.76; P, 3.43. Found: C, 62.79;
H, 10.18; N, 7.69; P, 3.41. ESI-MS, m/z: 301.1 (68), 884.6 (61)
[M þ H]þ, 906.7 (100) [M þ Na]þ. ESI-HRMS calcd for C47H91N5O8P
884.6600, found: 884.6606 [M þ H]þ; calcd for C47H90N5O8PNa
906.6419, found: 906.6425 [M þ Na]þ. 1H NMR (CDCl3 þ MeOD,
A mixture of 6-O-benzylpurine (9) or 2-amino-6-O-benzylpur-
ine (10) (20 mmol, 4.52 g or 4.82 g, respectively), S-(ꢀ)-glycidol
trityl ether (6.33 g, 20 mmol), Cs2CO3 (800 mg, 2.5 mmol) in DMF
(100 ml) was heated to 100 ꢁC for 10 h. The solution was evaporated
and the syrupy residue was chromatographed on a silica gel column
(500 g) in EtOAc/toluene (1:3).
ppm) d
: 0.85 (m, 6H, H-230), 1.18e1.32 (m, 52H, H-100 to H-220), 1.52
(m, 4H, H-90), 1.89 (m, 4H, H-60), 3.38 (m, 4H, H-80), 3.47 (m, 4H, H-
70), 3.56 (m, 2H, H-30), 3.76 (m, 1H, H-20), 3.85e3.96 (m, 2H, H-40),
4.10e4.18 (m, 5H, H-10b, H-50), 4.25 (m, 1H, H-10a), 7.74 (s, 1H, H-8).
4.4.1. 9-(S)-(2-Hydroxy-3-trityloxypropyl)-6-O-benzylpurine (11)
Yield 4.60 g (42%). Crystallization from toluene. Mp 180 ꢁC. Anal.
Calcd for C34H30N4O3: C, 75.26; H, 5.57; N,10.33. Found: C, 75.34; H,
5.62; N, 10.15.
13C NMR (CDCl3 þ MeOD, ppm)
d
: 13.40 (C-230), 22.17 (C-220), 25.64
(C-100), 28.87e29.20 (m, C-90, C-110 to C-200), 30.22 (d, J6 eP ¼ 5.9, C-
0
60), 31.44 (C-210), 43.07 (C-10), 59.54 (C-30), 62.35e63.87 (m, C-40, C-
50), 65.93 (C-70), 70.81 (C-80), 80.66 (d, J2 eP ¼ 10.9, C-20), 115.00 (C-
0
4.4.2. 9-(S)-(2-Hydroxy-3-trityloxypropyl)-2-amino-6-O-
benzylpurine (12)
5), 138.30 (C-8), 151.09 (C-4), 153.52 (C-2), 157.01 (C-6).
Yield 4.89 g (44%). Crystallization from toluene. Mp 177 ꢁC, Anal.
Calcd for C34H31N5O3: C, 73.23; H, 5.60; N, 12.56. Found: C, 73.15; H,
5.62; N, 12.46.
4.6. General method F
To a solution of 13 or 14 (0.5 mmol, 435 mg or 442 mg,
respectively) in dioxane (25 ml) an aqueous solution of NaOH (2M,
0.5 ml, 1.0 mmol) was added and the mixture was stirred for 48 h.
The solution was neutralized with hydrochloric acid (1 M, 1.1 ml,
1.1 mmol). Mixture was evaporated, codistilled with toluene and
the residue was chromatographed on a silica gel column (20 g) in
methanol/chloroform (1:4 / 1:2).
4.5. General method E
A mixture of 11 or 12 (2.4 mmol, 1.30 g or 1.34 g respectively)
and 8 (2.66 g, 3.6 mmol) in anhydrous DMF (20 ml) was treated
with NaH (60% suspension in mineral oil, 290 mg, 7.2 mmol) while
cooling down (ꢀ25 ꢁC). The mixture was stirred at a room
temperature overnight and then heated to 35 ꢁC for 8 h. The
mixture was then neutralized with acetic acid (0.27 ml, 4.8 mmol)
and partitioned between chloroform (100 ml) and aqueous satu-
rated solution of NaCl (100 ml). The aqueous portion was extracted
with chloroform again (50 ml). The combined organic portions
were dried with MgSO4 evaporated and the syrupy residue
was chromatographed on a silica gel column (200 g) in gradient
EtOAc/toluene (1:1) / EtOAc. Syrupy trityl derivative was dis-
solved in Et2O/MeOH (1:1, 10 ml). The solution was acidified with
methanolic solution of hydrogen chloride to pH 2e3 and stirred for
48 h. The product crystallized overnight in a fridge. The mother
liquors were concentrated and chromatographed on a silica gel
column in methanol/chloroform/hexane (1:5:5).
4.6.1. 3-(Hexadecyloxypropyl) ester of 9-(S)-[3-hydroxy-2-
(phosphonomethoxy)propyl]hypoxanthine (15)
Yield: 237 mg (81%). Mp 199e202 ꢁC. ESI-MS, m/z: 585.5 (100)
[M ꢀ H]ꢀ. ESI-HRMS calcd for C28H50N4O7P 585.3423, found:
585.3421 [M ꢀ H]ꢀ. HPLC purity 97.36% (254 nm). 1H NMR
0
0
0
(D2O þ NaOD, ppm) : 0.88 (t, 3H, J23 e22 ¼ 7.8, H-23 ),1.20e1.33 (m,
d
26H, H-100 to H-220),1.55 (m, 2H, H-90),1.86 (m, 2H, H-60), 3.38e3.74
(m, 8H, H-30, H-40, H-70, H-80), 3.91 (m, 2H, H-50), 4.15, 4.24 (m, m,
2H, H-10), 7.92 (s, 1H, H-2), 8.00 (s, 1H, H-8). 13C NMR (D2O þ NaOD,
ppm) d
: 14.47 (C-230), 23.22 (C-220), 26.61 (C-100), 29.97e30.55 (m,
C-90, C-110 to C-200), 31.22 (C-60), 32.54 (C-210), 44.28 (C-10), 60.87
(C-30), 62.80 (d, J5 eP ¼ 5.3, C-50), 65.67 (d, J4 eP ¼ 158.0, C-40), 67.91
0
0
(C-70), 71.52 (C-80), 80.98 (d, J2 eP ¼ 11.8, C-20), 123.44 (C-5), 141.38
0
(C-8), 150.39 (C-4), 154.03 (C-2), 168.18 (C-6).
4.5.1. Bis 3-(hexadecyloxypropyl) ester of 9-(S)-[3-hydroxy-2-
(phosphonomethoxy)propyl]hypoxanthine (13)
4.6.2. 3-(Hexadecyloxypropyl) ester of 9-(S)-[3-hydroxy-2-
(phosphonomethoxy)propyl]guanine (16)
Yield 1.27 g (61%). Mp 86 ꢁC. Anal. Calcd for C47H89N4O8P: C,
64.95; H, 10.32; N, 6.45; P, 3.56. Found: C, 64.99; H, 10.42; N, 6.44; P,
3.73; ESI-MS, m/z: 869.6 (24) [M þ H]þ, 891.7 (100) [M þ Na]þ.
Yield 265 mg (88%). Mp 235e238 ꢁC. Anal. Calcd for C28H54N5O8P
(monohydrate): C, 54.27; H, 8.78; N,11.30; P, 5.00. Found: C, 54.16; H,