Y. Hong et al.
The product-containing fractions were combined and lyophi-
Analytic HPLC condition: Column: BDS Hypersil C18,
lized to afford product 10 as a white powder (255 mg, 32%). 250 Â 4.6 mm, 5 mm; solvent: A, water with 0.1% TFA;
1
HRMS: calculated m/z = 482.16462 (M1Na) , observed B, acetonitrile with 0.1% TFA; flow: 1 mL/min; UV = 254 nm;
1
m/z = 482.16467 (M1Na) ; H NMR (DMSO-d
1
1
6
, 300.13 MHz):
.10 (dddd, 2H, J = 3.3, 12.8, 12.8, 12.8 Hz), 1.36 (dddd, 2H, J = 3.3,
T
R
= 8.7 min; 30% B isocratic.
Fractions containing pure product were pooled and concen-
1
2.8, 12.8, 12.8 Hz), 1.52 (m, 1H), 1.91 (m, 4H), 2.25 (dddd, 1H, trated by rotary evaporation. The final product was reconsti-
J = 3.4, 3.4, 12.2, 12.2 Hz), 2.36 (d, 2H), 3.58 (s, 3H), 4.28 (dd, 1H, tuted in ethanol to afford 170 mCi (56% yield) of 1a with a
J = 2.5, 4.5 Hz ), 4.41 (d, 1H), 4.46 (dd, 1H, J = 4.5, 6.5 Hz), 6.02 radiochemical purity 499%. The specific radioactivity was
1
3
(
d, 1H), 5.5–6.7 (b, 3H) 7.67 (bs, 2H), 8.50 (s, 1H) C NMR: 25.54, measured to be 48.07 Ci/mmol by LC-MS and tritium distribution
3
2
8
8.22, 30.98, 35.77, 42.05, 51.24, 73.30, 74.06, 81.12, 82.62, 85.65, of the compound was determined by H NMR (320 MHz,
3
6.77, 118.07, 139.87, 144.74, 149.51, 155.10, 171.91, 175.35.
3
d6-DMSO, T–H decoupled) d ppm 0.99–1.07 (methyl, H),
3 3
[
N-Ethyl-1,2- H]apadenoson, [(1R,4R)-methyl 4-(3-(6-amino-9- 3.20–3.30 (methylene, H), methyl/methylene H ratio, 5.58:1.
3
(
(2R,3S,4R,5S)-5-(1,2- H ethylcarbamoyl)-3,4-dihydroxytetrahydro-
furan-2-yl)-9H-purin-2-yl)prop-2-ynyl)cyclohexanecarboxylate]
1a): Compound 4 (300 mCi, 6.25 mmol) was dried under vacuum
in a 50-mL round-bottomed flask. Compound 10 (6 mg, The authors wish to thank Mr Robert Espina for the assistance in
3 mmol), (benzotriazol-1-yloxy)tripyrrolidino phosphonium hex- tritium NMR analysis and Mr Michael Lago for the assistance in
Acknowledgement
(
1
afluorophosphate (10 mg, 19.2 mmol), and DMF (5 mL) were GLP documentation.
added to the reaction flask at room temperature giving a clear
solution. Excess DIEA (0.2 mL) was then added and the solution
was stirred at room temperature. After 1.5 h HPLC indicated
References
3
complete consumption of the [ H]ethylamine and formation of
compound 1a. At this stage, the reaction was terminated by
addition of water (20 mL). The resulting solution was passed
through four Sep-Pak C18 cartridges (1 mL size) connected in
series. The cartridges were washed with 10 mL of water first and
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The ethanol fractions were combined and concentrated to a [4] S. Gabriel, Ber. 1887, 20, 2224.
[
[
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small volume for HPLC purification.
Preparative HPLC conditions: LUNA 5m C18(2) (10 Â 250);
solvent: A, water 1% TEAA; B, acetonitrile; flow: 4.7 mL/min;
UV = 254 nm; gradient: 0–35 min 25% B; 35–45 min 25–100% B.
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