title: A liquid chromatography and tandem mass spectrometry method for the determination of potential biomarkers of cardiovascular disease.
abstract: A simple, accurate and sensitive liquid chromatography tandem mass spectrometry (LC-MS/MS) method was developed and validated for the quantitation of α-ketoglutaric acid (α-KG), L-carnitine (L-CAR) and acetyl-L-carnitine (acetyl-L-CAR) in human urine as potential biomarkers of cardiovascular disease. The separation was performed using an isocratic elution of 0.1% formic acid in water and acetonitrile (97:3, v/v) on an Acclaim 120 C8 column (150 mm × 4.6 mm, 3.0 μm). The flow rate of the mobile phase was 1.2 mL/min and the total assay run time was 3 min. Detection was performed on a triple-quadrupole mass spectrometer in selected reaction monitoring (SRM) mode via an electrospray ionization (ESI) source in positive and negative ion modes. This method covered a linearity range of 0.1-500 ng/mL for L-CAR and acetyl-L-CAR and 1-1000 ng/mL for α-KG with lower limits of quantification (LLOQ) of 0.08 ng/mL for L-CAR, 0.04 ng/mL for acetyl-L-CAR and 0.8 ng/mL for α-KG. The intra-day and inter-day precision and accuracy of the quality control samples exhibited relative standard deviations of less than 5.54% and relative error values from -5.95% to 3.11%. Analyte stability was evaluated under various sample preparation, analysis and storage conditions and varied from -9.89% to -0.47%. A two-step solid-phase extraction (SPE) procedure using silica gel and quaternary amine cartridges was used for urine sample cleanup. The average recoveries for all analyzed compounds were better than 86.64% at three concentrations. The method was successfully applied for the quantitation of α-KG, L-CAR and acetyl-L-CAR in human urine samples.
authors: Department of Analytical Chemistry, Silesian University of Technology, 7 M. Strzody Str., 44-100 Gliwice, Poland. sylwia.magiera@polsl.pl
authors: Sylwia
authors: S
authors: Magiera
authors: Irena
authors: I
authors: Baranowska
authors: Jacek
authors: J
authors: Kusa
authors: Jacek
authors: J
authors: Baranowski
mesh_terms: D000108:Acetylcarnitine / Q000737:chemistry / Q000652:urine; D015415:Biomarkers / Q000737:chemistry / Q000652:urine; D002331:Carnitine / Q000737:chemistry / Q000652:urine; D016022:Case-Control Studies; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004355:Drug Stability; D006333:Heart Failure / Q000652:urine*; D006801:Humans; D007656:Ketoglutaric Acids / Q000737:chemistry / Q000652:urine; D016014:Linear Models; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D052616:Solid Phase Extraction; D021241:Spectrometry, Mass, Electrospray Ionization; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D015415:Biomarkers; D007656:Ketoglutaric Acids; D000108:Acetylcarnitine; D002331:Carnitine