Kinetics of enzymatic O-methylation: its value in assays for catecholamines in plasma

The kinetics of enzymatic O-methylation of catecholamines were studied under conditions like those used in the radioenzymatic assay of plasma catecholamines. Inappropriate Michaelis-Menten kinetics and linear approximations of exponential equations were not used. Mathematical analysis indicated the importance of the ratio of methyl donor (S-adenosylmethionine) to substrate (catecholamine) concentration. If the reaction is incomplete, only a large ratio will allow linear approximations between product formed and initial catecholamine concentration. The use of high-concentration internal standards to correct for plasma interference may give erroneous results by reducing this ratio. Accuracy will be improved by ensuring (a) that S-adenosylmethionine is always greatly in excess of catecholamine, (b) that concentrations of added standards are of the same order as for endogenous catecholamine, and (c) that a high activity of enzyme is used, to allow the reaction to reach completion even in the presence of some inhibition.