Quantification of monoamine biomarkers in cerebrospinal fluid: Comparison of a UHPLC-MS/MS method with a UHPLC coupled to fluorescence detection method.

Abstract

Inborn errors of monoamine neurotransmitter metabolism are rare genetic diseases classified as catecholamine and serotonin metabolism disorders or neurotransmitter transportopathies. To diagnose these orphan diseases, monoamine metabolites have been identified and validated as cerebrospinal fluid (CSF) biomarkers: 5-hydroxy-tryptophane, 5-hydroxy-indol-acetic acid, 3-ortho-methyl-DOPA, homovanillic acid, and 3-methoxy-4-hydroxyphenylglycol. The present work presents a UHPLC-MS/MS method developed for the quantification of these metabolites in CSF and compares it with a previously described UHPLC with fluorescence detection (UHPLC-FD) method. MS/MS detection was performed in positive electrospray ionization and multiple reaction monitoring mode. The UHPLC-MS/MS and UHPLC-FD methods were validated in terms of accuracy, linearity, precision and matrix effect. The lower limits of quantification (LLOQ) ranged between 0.5and 10nm and between 1 and 5nm for the UHPLC-MS/MS method and the UHPLC-FD one, respectively. We verified the applicability of both methods by analyzing 30 CSF samples. The measured concentrations were comparable with the reference values described in the literature. The two methods allowed pathological samples to be distinguished from healthy ones for clinical diagnosis. UHPLC-MS/MS and UHPLC-FD methods exhibited very close LLOQs. As the UHPLC-MS/MS method is more selective, it allows faster analysis with a run time of 6min per run vs. 10min for the UHPLC-FD method.