Utilization of a derivatization method with multidimensional liquid chromatography-mass spectrometry for the detection of neurotransmitters

Neurotransmitters are endogenous compounds located in the brain and can occur in other parts of the body at low concentrations. They are a challenging group of compounds when it comes to analysis. Compounds with amine functionality such as dopamine and serotonin are sensitive to high pH and light and can spontaneously oxidize and degrade. Since neurotransmitters exist at low levels, it can be difficult to achieve sufficient spectrometry data and clean chromatography. In order to improve data acquisition, derivatization and the use of multidimensional liquid chromatography (2D-LC) was evaluated. Dopamine, L-3,4-dihydroxyphenylalanine (L-DOPA), serotonin, tyrosine, tryptophan, and noradrenaline were the compounds of interest with tyramine included as an internal standard. Data was collected before and after derivatization to compare the resulting chromatography. Chromatography was completed using a 6x6 grid of methods with variables of organic solvent, elution pH, loading pH, and trapping column chemistry utilizing different elution column chemistries. Derivatization was examined with Dabsyl-Cl and Dansyl-Cl at pH 8.5, 9.5, 10.5, and 14. A final method was chosen with Dabsyl at pH 8.5 for the Multiple Reaction Monitoring (MRM) scans. Before derivatization, the chromatography had poor peak shapes such as tailing, fronting, or shouldering or too much distortion to be able to distinguish a peak for all of the examined methods. Resulting chromatography after derivatization showed overall improvement in peak shape and intensity for a majority of the methods. Derivatization aided in increasing the mass and stability of the compounds which allowed for more sensitive detection. Multidimensional liquid chromatography improved the separation of structurally similar compounds and increased sensitivity. By combining the two, better analysis of neurotransmitters was possible