Evaluation of a Liquid Chromatography–Tandem Mass Spectrometry Method for the Analysis of Glucosylceramide and Galactosylceramide Isoforms in Cerebrospinal Fluid of Parkinson’s Disease Patients

• Published in: Analytical chemistry (Washington) Vol. 96; no. 31; pp. 12875 - 12882
• Main Authors: Castillo-Ribelles, Laura, Arranz-Amo, Jose Antonio, Hernández-Vara, Jorge, Samaniego-Toro, Daniela, Enriquez-Calzada, Silvia, Pozo, Sara Lucas-Del, Camprodon-Gomez, Maria, Laguna, Ariadna, Gonzalo, Mercedes Arrúe, Ferrer, Roser, Martinez-Vicente, Marta, Carnicer-Caceres, Clara
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 06.08.2024
• Subjects: Bioaccumulation; Biological properties; Biological samples; Biomarkers; Biomarkers - cerebrospinal fluid; Cerebrospinal fluid; Chromatography; Chromatography, High Pressure Liquid; Galactosylceramide; Galactosylceramides - cerebrospinal fluid; Glucosylceramidase; Glucosylceramidase - cerebrospinal fluid; Glucosylceramidase - genetics; Glucosylceramides - cerebrospinal fluid; High performance liquid chromatography; Humans; Isoforms; Liquid chromatography; Mass spectrometry; Mass spectroscopy; Movement disorders; Mutation; Neurodegenerative diseases; Parkinson Disease - cerebrospinal fluid; Parkinson's disease; Performance evaluation; Risk factors; Scientific imaging; Sphingolipids; Substrates; Tandem Mass Spectrometry
• ISSN: 0003-2700; 1520-6882; 1520-6882
• DOI: 10.1021/acs.analchem.4c02654

Mutations in GBA1, encoding glucocerebrosidase beta 1 (GCase), are the most common genetic risk factor for Parkinson’s disease (PD). GCase dysfunction leads to an accumulation of glucosylceramide (GluCer) substrates in different organs and fluids. Despite the challenges in quantifying GluCer isoforms in biological samples, their potential clinical interest as PD biomarkers justifies the development of robust assays. An extensively evaluated high-performance liquid chromatography–tandem mass spectrometry (HPLC–MS/MS) method for quantifying 14 GluCer and galactosylceramide (GalCer) isoforms in human cerebrospinal fluid (CSF) samples is presented. Sample pretreatment, HPLC, and MS/MS parameters were optimized. Evaluation was performed according to the recommendations of the Clinical and Laboratory Standards Institute and European Medicines Agency guidelines. Four 7-point calibration curves were generated, with a linearity interval from 2.5 to 200 nM (R 2 ≥ 0.995). The limit of quantification was set at 5 nM. Between-run precision and accuracy were up to 12.5 and 9%, respectively. After method validation, we measured the levels of GluCer and GalCer isoforms in CSF human samples, including 6 healthy controls (HC), 22 idiopathic GBA1 wild-type PD (iPD) patients, and 5 GBA1-associated PD (PD-GBA) patients. GluCer/GalCer median ratios were found to be higher in the CSF of PD-GBA patients, particularly in severe GBA1 mutations, than those in iPD and HC. The observed trends in GluCer/GalCer ratios among groups provide novel information for the comprehensive analysis of sphingolipids as potential biomarkers of PD.