| There are many signaling molecules in the brain including L-amino acids, peptides, indoleamines, catecholamines and other small molecules. In most cases, samples generated in vivo, from microdialysis, sampling probes, or tissue extracts are analyzed for neurotransmitters by HPLC with electrochemical detection, radioimmunoassay, fluorometric detection, microelectrode, or fiber optic sensors. However, these traditional approaches are usually only capable of monitoring single neurotransmitters or a single class of neurotransmitters in a given experiment. A separation technique following sampling is attractive because it permits the simultaneous analysis of multiple analytes. Simultaneous monitoring of neurotransmitters is important to investigate the interactions and interconnections among different neurotransmitter systems, to correlate behavior, stimuli or drugs to neurotransmitter activity and to determine the role of each neurotransmitter.; The increased interest in stereochemical aspects of pharmacological activity and drug design has led to the development of new, specific and sensitive methods for the determination of enantiomers. Recent studies have provided evidence indicating that D amino acids, such as D-serine (Ser), D-aspartate (Asp) and D-alanine (Ala), present in mammals may have important functions. Thus, it has become important to be able to determine the D and L forms of amino acids in biological or physiological samples.; In this work, a sensitive and quantitative method for the simultaneous detection of various amine-containing neurotransmitters representing different classes was developed. The method is based on pre-column derivatization with o-pthaldialdehyde (OPA) and a thiol, gradient elution; on-column preconcentration (50 μm i.d. capillary columns) and detection by amperometry with a carbon fiber microelectrode. With this approach, simultaneous separation of L-amino acids, catechol and indole amines with concentration detection limits of less than 1 nM was achieved following injection of 250 nL samples. This represents the first example in which different classes of neurotransmitters are monitored simultaneously without the use of sample splitting. The method was extended to include amino acid enantiomers. Samples (100 nL) containing amino acid enantiomers, non-chiral amino acids, catechol and indole amines resulted in detection limits of 3–280 nM. There are no known reports of simultaneous analysis of L- and D-amino acids by amperometry following derivatization with OPA and a thiol. |
