反应详情

In a clean, dry 500 mL round bottom flask equipped with a stirring bar and under a stream of argon slowly add dry and finely powdered 3′-O-Me-Guanosine (1), (6 g, 20 mmol) to a mixture of trimethylphosphate ((OMe)3P) (50 mL) and phosphorous oxychloride (POCl3) (6 mL, 60 mmol) at 0° C. in small portions with continuous stirring under argon. The mixture was kept at 0-4° C. and allowed to stir at least 19 hrs. Diethyl ether (200 mL) was added to extract the excess phosphorous oxychloride and to simultaneously precipitate the 3′-O-methylguanosine-5′-phosphodichloridate, which was then pelleted by centrifugation and dissolved in 100 mL ice-cold 5% NaHCO3 in water. The resulting aqueous solution was adjusted to pH ˜1.5 using 1 N NaOH. After stirring at 0-4° C. for an additional 20 h, the pH was adjusted to 7.0 and the resulting mixture was applied to a column of DEAE Sephadex A25. The column was washed with 5 mM TEAB buffer, pH 7.5 and then eluted with freshly prepared 1M Triethylammonium bicarbonate (TEAB) buffer, pH 7.5. Fractions containing the 3′-O-Me GMP TEA salt (2) were pooled, concentrated to dryness.

1. customIn a clean, dry 500 mL round bottom flask
2. customequipped with a stirring bar and under a stream of argon
3. additionslowly add
4. customdry
5. customwas kept at 0-4° C.
6. stirringto stir at least 19 hrs
7. extractionto extract the excess phosphorous oxychloride
8. customto simultaneously precipitate the 3′-O-methylguanosine-5′-phosphodichloridate, which
9. dissolutiondissolved in 100 mL ice-cold 5% NaHCO3 in water
10. stirringAfter stirring at 0-4° C. for an additional 20 h
11. washThe column was washed with 5 mM TEAB buffer, pH 7.5
12. washeluted with freshly prepared 1M Triethylammonium bicarbonate (TEAB) buffer, pH 7.5
13. additionFractions containing the 3′-O-Me GMP TEA salt (2)
14. concentrationconcentrated to dryness