Dimethyl Sulfoxide → Methylenedioxyethamphetamine

HRID 1828979

Reaktionsgleichung

Reaktionsgleichung von HRID 1828979
CS(C)=O.O=C1CCC(=O)N1OC(=O)CCC(NC[CH:20]1[O:24][C:23]2[CH:25]=[CH:26][C:27]([CH2:29][CH:30]([N:32](CC)[C:33](=O)[C:34](F)(F)F)[CH3:31])=[CH:28][C:22]=2[O:21]1)=O.N[C@H](C(O)=O)CCCCN>P([O-])([O-])([O-])=O.[K+].[K+].[K+].CN(C=O)C>[CH3:34][CH2:33][NH:32][CH:30]([CH2:29][C:27]1[CH:26]=[CH:25][C:23]2[O:24][CH2:20][O:21][C:22]=2[CH:28]=1)[CH3:31]

Komponenten

6

Edukt 3

HCID 679Dimethyl SulfoxideHCID 5962L-LysineHCID 86595989N-(5-{2-[ethyl-(2,2,2-trifluoro-acetyl)-amino]-propyl}-benzo[1,3]dioxol-2-ylmethyl)-succinamic acid 2,5-dioxo-pyrrolidin-1-yl ester

Produkt 1

HCID 105039Methylenedioxyethamphetamine

Reagenzien, Katalysatoren, Lösungsmittel 2

HCID 6228DimethylformamideLösungsmittelHCID 62657Potassium Phosphate, TribasicLösungsmittel

Versuchsvorschrift

A solution of 188 mg of keyhole limpet hemocyanin (KLH) in 5.5 ml of 50 mM potassium phosphate (pH 7.5) was cooled in an ice-bath. To the solution were added 6 mL of dimethylsulfoxide dropwise, and the reaction temperature was maintained below room temperature. Then a solution of 54 mg (0.10 mmol) of 2L in 1.2 mL of DMF was added to the protein solution dropwise. The mixture was allowed to stir at room temperature 18 hours. The resulting conjugate was placed in a dialysis tube (10,000 MW cut-off) and was dialyzed in 1 L of 70% DMSO in 50 mM potassium phosphate (pH 7.5, 3 changes, at least 3 hours each), 1 L of 50% DMSO in 50 mM potassium phosphate (at least 3 hours), 1 L of 30% DMSO in 50 mM potassium phosphate (at least 3 hours), 1 L of 10% DMSO in 50 mM potassium phosphate (at least 3 hours) at room temperature. The trifluoroacetamido group of the conjugate was deprotected by dialysis of the resulting conjugate against 10% ammonium hydroxide for 3 days (1 L each for approximately 24 hours each), followed by 6 changes with 50 mM potassium phosphate (pH 7.5) at 4° C. (1 L each for at least 6 hours each). The protein concentration was determined to be 2.1 mg/mL using BioRad Coomassie blue protein assay (Bradford, M., Anal. Biochem. 72:248, 1976). A total of 34 mL of the conjugate was obtained. The extent of available lysine modification was determined to be 60% by the TNBS method (Habeeb AFSA, Anal. Biochem. 14:328-34, 1988).

Aufarbeitung

  1. temperaturethe reaction temperature was maintained below room temperature
  2. customThe resulting conjugate was placed in a dialysis tube
  3. custom(10,000 MW cut-off) and was dialyzed in 1 L of 70% DMSO in 50 mM potassium phosphate (pH 7.5, 3 changes, at least 3 hours each), 1 L of 50% DMSO in 50 mM potassium phosphate (at least 3 hours), 1 L of 30% DMSO in 50 mM potassium phosphate (at least 3 hours)
  4. custom1 L of 10% DMSO in 50 mM potassium phosphate (at least 3 hours)
  5. customat room temperature
  6. customThe trifluoroacetamido group of the conjugate was deprotected by dialysis of the resulting conjugate against 10% ammonium hydroxide for 3 days (1 L each for approximately 24 hours each),
  7. customfollowed by 6 changes with 50 mM potassium phosphate (pH 7.5) at 4° C.
  8. customfor at least 6 hours
  9. concentrationThe protein concentration
  10. customA total of 34 mL of the conjugate was obtained

Quellen und Nachweise

ORD-Eintrag
ord-f54ed04690a6404aa9f7ca35a30b0e94
Quelldatensatz
uspto-grants-2006_06
Patentnummer
US07060847B2

Verknüpfte Notizen

Wird geladen…