O-methylation of dopamine-0-sulfates with cathechol-0-methyltransferase

[²H₂]-dopamine-3-0-sulfate(DM-3-0-S) and [²H₂]-dopamine-4-0-sulfate (DM-4-0-S) were synthesized to investigate the possibility of their being substrates for catechol-0-methyltransferase (COMT). [²H₅]-3-0-methyldopamine (3-0-Me-DM) and [²H₅]-4-0-methyldopamine (4-0-Me-DM) were also synthesized as internal standards for the determination of enzymatic products by gas chromatography-mass spectrometry (GC-MS). [²H₂]-DM-3-0-S or [²H₂]-DM-4-0-S was incubated at 37° for 60 min in the presence of S-adenosyl-L-methionine with a crude enzyme preparation obtained from rat liver homogenate. The incubation mixture was treated with 0.5N HCl at 100°C for 1h to hydrolyze the remaining sulfate moiety. The reaction products were extracted with an Amberlite XAD-4, derivatized with pentafluoropropionic anhydride and determined by GC-MS. When [²H₂]-DM-3-0-S was used as a substrate, [²H₂]-3-0-Me-DM was found to be a major product accompanied by [²H₂]-4-0-Me-DM as a minor product. The ratio of [²H₂]-3-0-Me-DM to [²H₂]-4-0-Me-DM was found to be 26:1, while the ratio was 5.4:1 when [²H₂]-dopamine was used as a substrate. When [²H₂]-DM-4-0-S served as a substrate, [²H₂]-3-0-Me-DM was preferentially produced without detectable formation of [²H₂]-4-0-Me-DM.