Catecholamine analysis with microcolumn LC-peroxyoxalate chemiluminescence reaction detection

Abstract

Background Plasma catecholamines (CAs) are widely used as an index of sympathetic nervous system activity. In addition, CAs are known to be metabolized by catechol- O-methyltransferase (COMT) to produce their 3- O-methyl metabolites. We previously established a sensitive determination method of CAs and their 3- O-methyl metabolites using HPLC-peroxyoxalate chemiluminescence (POCL) reaction detection system. In this study, a microcolumn (100 × 1.0 mm I.D.) was used for separation to obtain higher sensitivity and shorter analysis time. Methods The system included automated precolumn ion-exchange extraction of amines, followed by separation on an ODS column, coulometric oxidation, fluorescence derivatization with ethylenediamine, and finally POCL reaction detection. Results The detection limits for CAs and their 3- O-methyl metabolites were 0.3–2.0 fmol. The analysis time was about 35 min, about half that of previously reported results. The method developed was used in monitoring changes in CAs and 3- O-methyl metabolite concentrations in human plasma during exercise. Conclusion The simultaneous determination method for concentrations of CAs and their 3- O-methyl metabolites in human plasma was developed using micro LC-peroxyoxalate chemiluminescence detection. We were successful in quantitating the changes in plasma CAs and their 3- O-methyl metabolites during exercise.