Simultaneous determination of monoamines, their precursor amino acids, and related metabolites in mice brain by high-performance liquid chromatography with a coulometric electrode array system

A method based on high performance liquid chromatography with a coulometric electrode array system (HPLC-coulometric electrode array) using C-18 column has been developed for the simultaneous determination of norepinephrine (NE), epinephrine (E), l-3,4-dihydroxyphenylalanine (l-DOPA), p-tyrosine (p-TYR), dopamine (DA), m-tyrosine (m-TYR), 5-hydroxytryptamine (5-HT), homovanillic acid (HVA), and 5-hydroxyindoleacetic acid (5-HIAA) in mice brain. The chromatography was performed using a C-18 column (250mm x 4.6mm i.d. and 5 mu m) with sodium acetate buffer (pH 5.0, 0.05 M) and methanol as the mobile phase. Elution of analytes was carried out at a flow rate of 1.0 ml/min. The nine compounds were monitored using an ESA electrochemical detector. Potentials of three electrodes in series were set at 200, 500, and 700 mV, respectively. Optimization of the pH of the mobile phase and the proportion of methanol were also considered. The minimal detection limits were 2-8 ng/ml. Linear (r = 0.99) detector performances were observed within a range of 10 similar to 2000 ng/ml. Recoveries for the nine compounds in spiked samples were over 90% and the relative standard deviations (RSD) were less than 4.0%.