Development of a nano-electrospray MSn method for the analysis of serotonin and related compounds in urine using a LTQ-orbitrap mass spectrometer.

Serotonin, its key metabolite hydroxyindole acetic acid (5-HIAA) and dopamine have been shown to be potential biomarkers whose levels in serum and urine can be correlated with certain psychiatric and physiological disorders and illness, including depression, schizophrenia, anxiety and dementia. Recently we have published elsewhere that 5-HIAA has been identified as a potential biomarker for Attention Deficit Hyperactivity/Hyperkinetic Disorder (AD-HKD). This study describes a versatile and validated method for the analysis of these three compounds in urine using a nanoelectrospray-MS(n) method interfaced with an LTQ Orbitrap mass spectrometer. No chromatographic separation is required prior to nanoelectrospray infusion. Good linear calibrations were obtained for analytes in urine (with serotonin and dopamine giving R(2)=0.9999 and 5-HIAA having a lower R(2) value of 0.9955). Acceptable intraday repeatability was achieved for all analytes with RSD values (n=5) ranging from 4.4% to 6.2% (57, 65 and 52 nmol/L for serotonin, dopamine and 5-HIAA respectively) to 2.1-8.1% (2837, 3268, 2618 nmol/L for serotonin, dopamine and 5-HIAA respectively). Excellent limits of detection (LOD) and limits of quantitation (LOQ) were achieved with spiked samples for all compounds; with LODs of 9-12.9 nmol/L and LOQs of 27.2-57.7 nmol/L for analytes in urine. An appropriate sample clean-up procedure for urine was developed to ensure highest recovery and reproducibility on analysis.