Assay of 3-nitrotyrosine in tissues and body fluids by liquid chromatography with tandem mass spectrometric detection

Abstract

3-Nitrotyrosine (3-NT) is a marker of protein nitration in physiological systems. It is present as 3-nitrotyrosine residues in proteins of tissue, extracellular matrix, plasma, and other body fluids and food. It is also present in body fluids and some beverages as free nitrotyrosine and is excreted in urine with the major urinary metabolite 3-nitro-4-hydroxyphenylacetic acid. Quantitation of 3-nitrotyrosine requires tandem mass spectrometry for specific detection. The method developed to determine 3-nitrotyrosine (along with protein glycation and oxidation adducts in a quantitative screening assay) by liquid chromatography with tandem mass spectrometric detection is described. The 3-NT residue contents of plasma protein, hemoglobin, lipoproteins, and cerebrospinal fluid protein and the concentrations of free 3-nitrotyrosine in plasma, urine, and cerebrospinal fluid are given. Changes of 3-nitrotyrosine residue and free 3-nitrotyrosine in diabetes, cirrhosis, acute and chronic renal failure, and neurological disorders, including Alzheimer's disease, are presented and compared with independent estimates.