Abstract
Urine represents a convenient biofluid for metabolomic studies due to its noninvasive collection and richness in metabolites. Here, amino acids are valuable biomarkers for their ability to reflect imbalances of different biochemical pathways. An impact of amino acids on pathology, prognosis and therapy of various diseases, including inflammatory bowel disease (IBD), is therefore the subject of current clinical research. This work is aimed to develop a capillary electrophoresis-tandem mass spectrometry (CE-MS/MS) method for the quantification of the 20 proteinogenic amino acids in human urine samples obtained from patients suffering from IBD and treated with thiopurines. The optimized CE-MS/MS method, with minimum sample preparation (just “dilute and shoot”), exhibited excellent linearity for all the analytes (coefficients of determination were higher than 0.99), with inter-day and intra-day precision yielding relative standard deviations in the range of 0.91–15.12% and with accuracy yielding relative errors in the range of 85.47–112.46%. Total analysis time, an important parameter for the sample throughput demanded in routine practice, was shorter in ca. 17% when compared to established CE-MS methods. Favorable performance of the proposed CE-MS/MS method was also confirmed by the comparison with corresponding ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS) method. Consistent data for the investigated amino acid metabolome were obtained using both methods. For the first time, the amino acid profiling by CE-MS approach was applied on the clinical IBD samples. Here, significant differences observed in the concentration levels of some amino acids between IBD patients undergoing thiopurine treatment and healthy volunteers could result from the simultaneous action of the disease and the corresponding therapy. These findings indicate that amino acids analysis could be a valuable tool for the study of mechanism of the IBD treatment by thiopurines.
Highlights
Qualitative and/or quantitative measurement of targeted compounds of metabolism in human body fluids has been established as a crucial tool for the prediction, diagnosis or investigation of the mechanism of various diseases [1,2,3,4]
Favorable performance and reliability of the proposed Capillary electrophoresis (CE)-MS/MS method, confirmed by the comparative UHPLC-MS analysis, are attributes demanded in routine clinical analysis
The both developed methods reflected in the successful profiling of the clinical samples obtained from 13 inflammatory bowel disease (IBD) patients
Summary
Qualitative and/or quantitative measurement of targeted compounds of metabolism in human body fluids has been established as a crucial tool for the prediction, diagnosis or investigation of the mechanism of various diseases [1,2,3,4]. Amino acids play an important role in cell metabolism and are associated with different functions—some act as neurotransmitters, serve as essential precursors for the synthesis of variety of molecules with enormous importance or regulate key metabolic pathways and processes that are vital to the health, growth, development, reproduction and homeostasis of organisms [5]. Disturbances in amino acids levels may lead to different metabolic diseases or may indicate disease status. The role of amino acids in origin, progression or control of different diseases is widely studied [6]. Amino acids are important for intestinal growth, mucosal integrity and serve as the precursors of metabolically active proteins, glutathione, nitric oxide or polyamines [12], and, they may have some impact on the IBD status. Vidal-Lletjós et al [13] reported the study on the influence of glutamine and arginine on IBD progression
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