764
Elbert, Břehová, Holý:
Jgem = 13.9, J(H-C-P) = 8.9, 1 H and 3.56 dd, Jgem = 13.9, J(H-C-P) = 9.3, 1 H (PCH2O);
1.17 d, J(3’-2’) = 6.4, 3 H (H-3’); 1.16 d, 3 H, 1.11 d, 3 H, 1.08 d, 3 H, 1.03 d, 3 H,
J(CH3-CH) = 6.2 (CH3 ipr.). 13C NMR (DMSO-d6): 154.99 (C-6); 152.94 (C-2); 151.19 (C-4);
127.17 (C-8); 119.17 (C-5); 74.98 d, J(2’-P) = 12.0 (C-2’); 70.35 d, J(C-O-P) = 6.4 and 70.22 d,
J(C-O-P) = 6.3 (CH ipr.); 62.72 d, J(C-P) = 164.4 (PCH2O); 48.69 (C-1’); 23.65–23.99 m (CH3
ipr.); 16.94 (C-3’).
8-Bromo-(R)-9-(2-phosphonomethoxypropyl)adenine (9)
Diester 8 (540 mg, 1.2 mmol) in acetonitrile (10 ml) was treated with Me3SiBr (1 ml) at
room temperature overnight. The volatiles were removed under reduced pressure and the
residue was co-distilled with water. The crude product was purified by the preparative HPLC
(a gradient of MeOH (0–100%) in H2O). Crystallization from ethanol afforded a white crys-
talline product (150 mg, 35%), whose m.p. is not reached below 250 °C, decomposition.
ESI MS: 364.1 (38) [M – H]–. 1H NMR (D2O, ref. dioxane): 8.37 s, 1 H (H-2); 4.37 m, 2 H
(H-1’); 4.05–4.12 m, 1 H (H-2’); 3.62 dd, 1 H, J(H-C-P) = 8.90, Jgem = 13.48 and 3.43 dd,
1 H, J(H-C-P) = 8.90, Jgem = 13.47 (OCH2P); 1.28 d, 3 H, J(3’-2’) = 6.29 (H-3’). 13C NMR
(D2O, ref. dioxane): 150.92 (C-6); 150.09 (C-4); 145.95 (C-2); 132.53 (C-8); 119.44 (C-5);
76.63 d, J(2’-P) = 10.27 (C-2’); 65.62 d, J(CH2-P) = 157.73 (OCH2P); 50.31 (C-1’); 17.14
(C-3’). For C9H13BrN5O4P (366.11) calculated: 29.53% C, 3.58% H, 21.83% Br, 19.13% N,
8.46% P; found: 29.36% C, 3.62% H, 21.57% Br, 18.90% N, 8.00% P.
9-(2-Phosphonomethoxyethyl)-2,6-diamino-[8-3H]purine (4)
To 8-bromo PMEDAP 1 (6 mg, 16.3 µmol) in the presence of 27 mg 5% PdO on BaSO4 in a
small flask equipped with a magnetic stirrer, water (1.2 ml) and triethyl amine (0.13 ml)
were added. The reaction mixture was degassed by three successive freeze-thaw cycles under
vacuum. 5 Ci of carrier-free 3H2 were transferred to the reaction flask (with a starting pres-
sure of 640 torr) and the reaction was left to proceed under vigorous agitation at room tem-
perature for 1.5 h. The residual tritium gas was pumped off and the reaction mixture was
transferred to a heart-shaped flask (the reaction flask was washed 3 times with 1 ml of wa-
ter, added to the reaction mixture). Acetic acid (0.5 ml) was added and the solvents were
evaporated in a closed system under vacuum (the solvent trap was cooled by liquid nitro-
gen). The residue was dispersed in 3 ml of water and the mixture was evaporated to dryness.
This procedure was repeated once more. After the last evaporation, the residue was dispersed
in 3 ml of water and the catalyst was separated by centrifugation. The supernatant was
taken off and the catalyst was washed three times with 2 ml of water. The volume of the
unified supernatants was adjusted to 10 ml with water. The total activity of the crude prod-
uct was 25.3 mCi. According to the HPLC analysis of the crude product, there was no start-
ing compound and more than 95% of the activity was in desired product 4. The crude
product was purified on a semi-preparative Synergi 4µ Fusion-RP 80, 250 × 10 mm column
(Phenomenex, USA) in the isocratic mode with water containing 0.25% (v/v) of acetic acid
as the mobile phase. The yield was 17.65 mCi of the product with a radiochemical purity of
>99% (radio HPLC). The 3H NMR (D2O, ref. TDO) was 7.98 s (3H-8). The 1H NMR was
identical to that of the standard with the exception of the integration of the H-8 signal.
From the decrease of the intensity of the 1H-8 signal, the specific activity was calculated as
10.7 Ci/mmol. The UV method of specific activity assay consisted of a measurement of the
volume activity by LSC and an assay of the mass concentration by HPLC at 284 nm. The re-
Collect. Czech. Chem. Commun. 2010, Vol. 75, No. 7, pp. 757–766