Determination of picogram levels of brain catecholamines and indoles by a simplified liquid chromatographic electrochemical detection method

Abstract

A simple and rapid method for the simultaneous determination of norepinephrine, epinephrine, dopamine, 5-hydroxytryptamine and 5-hydroxyindoleacetic acid in rat brain regions by high-performance liquid chromatography (HPLC) with electrochemical detection has been developed. Perchloric aced extracts of the tissue were directly analyzed in the HPLC system. Each of these compounds gave a linear response over the range of 10–320 ng/ml cerebellar homogenate (0.2–6.4 ng on column). Analytical recoveries of these compounds added to the homogenates were complete when compared with standards dissolved in perchloric acid. The average between-run coefficients of variation for all these compounds were lower than 6.7%, over the range of 10–320 ng/ml, whereas the within-run coefficients of variation at 10 ng/ml were lower than 6.9%. Under the present instrumental and mobile phase conditions, all compounds were readily oxidized at 0.72 V vs. a Ag/AgCl reference electrode. The present method has been applied to a study determining the basal levels of these compounds in several rat brain regions as well as levels after medium raphe lesions.