Determination of catecholamines by radioenzymatic assay using ion-pair liquid chromatography

A simple radioenzymatic method for the simultaneous determination of norepinephrine, epinephrine and dopamine in human serum was developed. catecholamines were converted to normetanephrine (NMN), metanephrine (MN), 3-methoxytyramine (3-MT) and their 4-O-methyl isomers by catechol O-methyl transferase with S-adenosylmethionine-(3H-methyl) ([3H]SAM). After addition of unlabelled NMN, MN and 3-MT as carriers, the methyl derivatives were isolated from the reaction mixture by passing through a small boric acid gel column and separated from each other by ion-pair liquid chromatography. Interference by tailing of radioactivity of [3H] SAM and its degradation products eluted earlier was minimized by the use of a precolumn and a branched flow path.