Cortisol quantification in human plasma and urine by liquid chromatography coupled to mass spectrometry: Validation, analysis and application in a reference population and patients with adrenal incidentalomas
Andressa Cristina dos Santos Marques , Bruna Brito , Jéssica Gorett Brito Fontes , Gabriel Reis Alves Carneiro , João Felipe Dickson Rebelo , Aline Barbosa Moraes , Leonardo Vieira Neto , Monica Costa Padilha
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引用次数: 0
Abstract
Cortisol is a glucocorticoid hormone, which is involved in cardiovascular, metabolic, inflammatory and behavioral processes in the human body. Immunoassays, used for routine analysis of analytes, usually lead to erroneous quantification due to the low specificity of these methods. In this study, we developed LC-MS/MS methods with surrogating matrices for evaluating cortisol in both urine and plasma samples and validated them according to Brazilian Health Regulatory Agency guidelines. Urine samples were prepared with an enzymatic hydrolysis stage, followed by a solid-phase extraction (C18 cartridges) using dichloromethane:methanol 9:1 as elution solvent. Plasma samples were prepared by protein precipitation with acetonitrile, using 50 μL of sample. HPLC was performed using a C8 column under 300 mL min−1 flow gradient conditions with water and methanol, both containing 5 mM ammonium formate and 0.1 % formic acid. Mass spectrometer with electrospray ionization in positive mode and selected reaction monitoring as detection technique were employed. Calibration curves were linear over a concentration range of 1–200 ng mL-1 for urine (r2 = 0.9950) and 0.5–300 ng mL-1 for plasma (r2 = 0.9970). The methods were selective, showed suitable precision, accuracy, and sensibility (limit of quantification = 0.85 ng mL−1 for urine and 0.15 ng mL−1 for plasma). Validated methods were successful applied to 22 real samples and a cohort of patients [n = 63 urines and n = 79 plasmas (from the Clementino Fraga Filho University Hospital, Rio de Janeiro, Brazil)].
期刊介绍:
The Official Journal of the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC)
Clinica Chimica Acta is a high-quality journal which publishes original Research Communications in the field of clinical chemistry and laboratory medicine, defined as the diagnostic application of chemistry, biochemistry, immunochemistry, biochemical aspects of hematology, toxicology, and molecular biology to the study of human disease in body fluids and cells.
The objective of the journal is to publish novel information leading to a better understanding of biological mechanisms of human diseases, their prevention, diagnosis, and patient management. Reports of an applied clinical character are also welcome. Papers concerned with normal metabolic processes or with constituents of normal cells or body fluids, such as reports of experimental or clinical studies in animals, are only considered when they are clearly and directly relevant to human disease. Evaluation of commercial products have a low priority for publication, unless they are novel or represent a technological breakthrough. Studies dealing with effects of drugs and natural products and studies dealing with the redox status in various diseases are not within the journal''s scope. Development and evaluation of novel analytical methodologies where applicable to diagnostic clinical chemistry and laboratory medicine, including point-of-care testing, and topics on laboratory management and informatics will also be considered. Studies focused on emerging diagnostic technologies and (big) data analysis procedures including digitalization, mobile Health, and artificial Intelligence applied to Laboratory Medicine are also of interest.