Capillary electrophoresis tandem mass spectrometry determination of glutamic acid and homocysteine’s metabolites: Potential biomarkers of amyotrophic lateral sclerosis

Abstract

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease that affects both lower and upper motor neurons, leading to muscle atrophy, paralysis, and death caused by respiratory failure or infectious complications. Altered levels of homocysteine, cysteine, methionine, and glutamic acid have been observed in plasma of ALS patients. In this context, a method for determination of these potential biomarkers in plasma by capillary electrophoresis tandem mass spectrometry (CE-MS/MS) is proposed herein. Sample preparation was carefully investigated, since sulfur-containing amino acids may interact with plasma proteins. Owing to the non-thiol sulfur atom in methionine, it was necessary to split sample preparation into two methods: i) determination of homocysteine and cysteine as S-acetyl amino acids; ii) determination of glutamic acid and methionine. All amino acids were separated within 25min by CE-MS/MS using 5molL acetic acid as background electrolyte and 5mmolL acetic acid in 50% methanol/HO (v/v) as sheath liquid. The proposed CE-MS/MS method was validated, presenting RSD values below 6% and 11% for intra- and inter-day precision, respectively, for the middle concentration level within the linear range. The limits of detection ranged from 35 (homocysteine) to 268nmolL (glutamic acid). The validated method was applied to the analysis of plasma samples from a group of healthy individuals and patients with ALS, showing the potential of glutamic acid and homocysteine metabolites as biomarkers of ALS.

Recent advances in capillary electrophoresis‐mass spectrometry: Instrumentation, methodology and applications

“…Biomarker identification is performed in different biological fluids, blood being one of the most promising amongst them. In recent studies, metabolites and especially amino acids were identified as biomarkers in blood for amyotrophic lateral sclerosis , chronic kidney disease (CKD) , major depressive disorder , onchocercias infection , osteoporosis , Parkinson's disease , pancreatic cancer , urinary tract symptoms and metabolic syndrome in postmenopausal women by CZE‐MS. For identification and accurate quantification of biomarkers for inborn errors of metabolism from dried blood spots, DiBattista et al.…”

Recent advances in amino acid analysis by capillary electromigration methods: June 2015–May 2017

“…The separation occurred in less than 25 min using a 5 M acetic acid migration buffer with estimated LODs ranging from 35 nM (Hcy) to 268 nM (Glu). Analyses of plasma from patient with amyotrophic lateral sclerosis were described, demonstrating the potential of glutamic acid and homocysteine metabolites as biomarkers .…”

Circulating Metabolites as Potential Biomarkers for Neurological Disorders—Metabolites in Neurological Disorders

“…A study of blood plasma from ALS patients and healthy controls, using MS-targeted metabolomics, identified significantly higher levels of glutamic acid and homocysteine in the plasma of these patients [ 17 , 83 ]. Although those molecules are potential biomarkers for ALS, further studies are necessary to confirm their mechanisms for ALS physiology [ 83 ].…”

“…In addition, it is also possible to observe the involvement of harmful oxidative processes, with the potential to affect membranes, as has already been proposed for Lysophosphatidic acid C18: 2 in patients with AD [ 71 ]. Furthermore, carnitine and the related metabolite acetylcarnitine, an intermediary of fatty acid oxidation, were found in studies on AD, ALS, MS, and stroke and related to impairment mitochondrial and energy metabolisms and fatty acids transport during the catabolism of lipids [ 9 , 68 , 83 , 109 ]. Because of the acetyl group, the acetylcarnitine can cross the blood–brain barrier more easily and is the preferred form in the central nervous system [ 9 ].…”