Simultaneous measurement of serotonin, catecholamines and their metabolites in mouse brain homogenates by high-performance liquid chromatography with a microbore column and dual electrochemical detection.

A dual electrochemical detector with two working electrodes (anode and cathode) suitable for high-performance liquid chromatography with a microbore octadecylsilica column was applied for the simultaneous measurement of norepinephrine, epinephrine, dopamine, 3,4-dihydroxyphenylacetic acid, homovanillic acid, 5-hydroxyindoleacetic acid, 3-methoxytyramine and 5-hydroxytryptamine (serotonin) in mouse brain homogenates. Microbore high-performance liquid chromatography provides very good resolution of these analytes and offers selective detection of biogenic amines and their metabolites on the basis of their retention behaviour and electrochemical reversibility. The large early-eluting peak of brain homogenates was eliminated on cathodic detection, thereby providing reliable measurements of early eluates. The detection limit of this method was ca. 0.2-0.5 pg per injection for all components, at a signal-to-noise ratio of 3. Owing to the high sensitivity, the brain tissue samples could be kept very small (less than 10 mg). Isocratic separation of these analytes was achieved within 15 min; hence over 90 analyses could be performed in a single working day. This simple, efficient and sensitive method can be used as a basic research tool for the assaying of biogenic amines and their metabolites in brain homogenates.