Determination of dopamine-beta-hydroxylase in cerebrospinal fluid by high-performance liquid chromatography with...

An improved method is described for the measurement of dopamine-beta-hydroxylase (D beta H) activity in cerebrospinal fluid, which is based on an incubation with dopamine at a saturated substrate concentration and quantitation of the reaction product noradrenaline, by high-performance liquid chromatography with electrochemical detection using 3,4-dihydroxynorephedrine as internal standard. Sample workup consists in an ion pair extraction to isolate the catecholamines from the rather complex incubation medium, a cation ion exchange to eliminate the bulk amount of dopamine, and alumina adsorption to concentrate the sample prior to high-performance liquid chromatography. The methodology was used to evaluate some of the characteristics of D beta H in cerebrospinal fluid and the stability of the enzyme. The procedure was also employed to determine the change in the D beta H following drug administration. Intravenously administered yohimbine caused an increase in D beta H activity in cerebrospinal fluid of rabbits as expected from its known alpha 2-antagonist properties.