Simultaneous detection and quantification of 3-nitrotyrosine and 3-bromotyrosine in human urine by stable isotope dilution liquid chromatography tandem mass spectrometry.
Nitration and bromination of proteins, giving rise to the respective 3-nitrotyrosine (3NT) and 3-bromotyrosine (3BT), are implicated in asthma, allergic inflammatory disorders, and cancer. We have developed an isotope dilution liquid chromatography electrospray ionization tandem mass spectrometry (LC/MS/MS) assay for simultaneous analysis of protein-bound 3NT and 3BT in human urine. The detection limits (S/N=3) were 10 pg (44 fmol) for 3NT and 5.0 pg (19 fmol) for 3BT injected on-column. The average levels of protein-bound 3NT and 3BT in 23 healthy individuals were 9.7 +/- 11.0 (mean +/- S.D.) in 10(5) tyrosine and 4.4 +/- 3.9 (mean +/- S.D.) in 10(3) tyrosine, respectively, using this highly sensitive LC/MS/MS under the selective reaction monitoring mode. Furthermore, the levels of urinary 3NT and 3BT show a statistically significant correlation (R(2) = 0.55, p = 0.0065, n=23). The high specificity and accuracy of this LC/MS/MS method render it a valuable tool in measurement of 3NT and 3BrT in the human urinary protein as promising noninvasive biomarkers for protein tyrosine nitration and bromination in vivo.
Chen
Hauh-Jyun Candy
HJ
Department of Chemistry and Biochemistry, National Chung Cheng University, 168 University Road, Ming-Hsiung, Chia-Yi 62142, Taiwan. chehjc@ccu.edu.tw
Chiu
Wei-Loong
WL
D002138:Calibration; D002853:Chromatography, Liquid; D006801:Humans; D011865:Radioisotope Dilution Technique; D012680:Sensitivity and Specificity; D053719:Tandem Mass Spectrometry; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000652:urine
C002744:3-nitrotyrosine; D014443:Tyrosine; C118327:3-bromotyrosine