Separation of catecholamines and dopamine-derived DNA adduct using a microfluidic device with electrochemical detection

摘要

Dopamine (DA) can produce quinones that form protein and depurinating DNA adducts; the latter are thought to be involved in some neurode-generative diseases (e.g. Parkinson's disease) [E.L. Cavalieri et al., Carcinogenesis 23 (2002) 1071-1077]. Methods with high resolving power are needed to detect neurotransmitters (e.g. DA) and DA-derived DNA adducts in human fluids, as their presence is difficult to determine by standard chromatography with UV absorbance detection. We propose using microfluidic devices with an electrochemical (EC) detection system to determine the presence of DA, DA-6-A7Gua adduct, L-tyrosine, dihydroxyphenylalanine (L-DOPA), and catechol (used as an internal standard). A PDMS glass-based, hybrid microfluidic device for free solution electrophoresis with totally integrated electrodes as well as improved and simplified gated injection was fabricated and tested. The best position and optimal width for minimizing background current of the working electrode was experimentally established. Using a single power supply simplified the operation of the microfluidic device and provided control of the plug size (length), leading to increased separation quality. It is shown that excellent separation of the above analytes can be accomplished in a relatively short time; the response is linear in the range between 2 and 300 μM, and reproducible with a limit of detection (LOD) for DA-6-N7Gua adducts in the sub-femtomole range. Thus, future separation and identification of various neurotransmitters and/or their products in human fluids could be accomplished using microfluidic devices with electrochemical detection.