Trimethyl Phosphate → 3'-O-Methylguanosine 5'-(dihydrogen phosphate)

HRID 1873940Notes

Reaction equation

HRID 1873940 reaction equation
[P:1]([O:9][CH2:10][C@H:11]1[O:15][C@@H:14]([N:16]2[C:26]3[N:25]=[C:23]([NH2:24])[NH:22][C:20](=[O:21])[C:19]=3[N:18]=[CH:17]2)[C@H:13]([OH:27])[C@@H:12]1[OH:28])([O:4]P(O)(O)=O)(=[O:3])[OH:2].[CH3:29]OP(OC)(OC)=O.P(Cl)(Cl)(Cl)=O>C(OCC)C>[P:1]([O:9][CH2:10][C@H:11]1[O:15][C@@H:14]([N:16]2[C:26]3[N:25]=[C:23]([NH2:24])[NH:22][C:20](=[O:21])[C:19]=3[N:18]=[CH:17]2)[C@H:13]([OH:27])[C@@H:12]1[O:28][CH3:29])([OH:4])([OH:2])=[O:3]

Participants

5

Reactant 3

HCID 10541Trimethyl PhosphateHCID 24813Phosphorus oxychlorideHCID 135398619Guanosine Diphosphate

Product 1

HCID 1362505513'-O-Methylguanosine 5'-(dihydrogen phosphate)

Reagents, catalysts & solvents 1

HCID 3283Diethyl EtherSolvent

Procedure

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.

Workup

  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

Sources & evidence

ORD record
ord-336ac4ba64214720be1c8ad4df49f818
Source dataset
uspto-grants-2012_11
Patent number
US08304529B2

Reaction notes

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