Reassessment of the antioxidative mixture for the challenging electrochemical determination of dopamine, noradrenaline and serotonin in microdialysis samples

• Monoamine monitoring by in vivo microdialysis is a valuable research tool.
• Simultaneous analysis of monoamines using (U)HPLC–ECD remains challenging.
• Addition of antioxidative agents interferes with the analysis of noradrenaline.
• Monoamine stability in different antioxidative mixtures is evaluated.
• The presented mixture offers opportunities for further analytical improvements.

In recent years, the simultaneous monitoring of the monoamine neurotransmitters dopamine, noradrenaline and serotonin in vivo is advancing due to innovations in miniaturized and fast chromatographic techniques. However, the determination of the most hydrophilic compound, noradrenaline, in microdialysis samples by (ultra-)high performance liquid chromatography ((U)HPLC) with electrochemical detection (ECD) is impeded by a broad solvent front, caused by the addition of antioxidative agents. Hence, an elaborate reassessment of currently used antioxidative mixtures is necessary for further analytical improvements. The proposed mixture, containing 100 mM acetic acid, 0.27 mM Na2EDTA and 12.5 μM ascorbic acid (pH 3.2), is less complex than previously described mixtures and shows minimal ECD interference. It stabilizes the three monoamines in standard solutions and in microdialysis samples, considering both autosampler stability at 4 °C for 48 h and long term stability at −20 °C for a duration of six months. An in vivo microdialysis experiment demonstrates the possibility to monitor changes in extracellular levels of the three monoamines simultaneously in the rat hippocampus with UHPLC–ECD using the optimized antioxidative mixture.