Ultrasensitive peroxynitrite-based luminescence with L-012 as a screening system for antioxidative/antinitrating substances, e.g. Tylenol (acetaminophen), 4-OH tempol, quercetin and carboxy-PTIO.
Previously our group developed a water-soluble antioxidant screening system using the luminescence of the reaction of peroxynitrite and luminol. In the present study we replaced luminol with the luminol-like compound L-012. This increases the production of luminescence approximately 100-fold and therefore, with a higher signal:noise ratio, this new system can detect antioxidation and antinitration effects at lower doses of the inhibitor. We studied acetaminophen (Tylenol) and its metabolite 3-nitroacetaminophen, tyrosine and nitrotyrosine and all these substances were inhibitory in a dose-responsive manner and below micromolar amounts. In addition quercetin, a polyphenol, was highly active (below micromolar amounts) as an antioxidant and antinitrating compound. 4-OH tempol, the stable free radical, superoxide dismutase (SOD) mimetic, was inhibitory in a dose-responsive manner and below micromolar amounts. Carboxy-PTIO was inhibitory at 10 times micromolar amount but not below that dose, which may be related to colour quenching, since the drug is deeply blue, or possibly it is an inhibitor with a slow kinetic profile. Finally, the amino acid tyrosine has been found to be inhibitory in micromolar amounts, similar to acetaminophen. This indicates that tyrosine can act as an antioxidant and antinitration target alone or conjugated in protein, e.g. insulin.
Van Dyke
Knox
K
Department of Biochemistry and Molecular Pharmacology, Robert C. Byrd Medical Center, West Virginia University Medical School, Morgantown, WV, USA. kvandyke@hsc.wvu.edu
Ghareeb
Erica
E
Van Dyke
Mark
M
Van Thiel
David H
DH
D000082:Acetaminophen; D000975:Antioxidants / Q000032:analysis*; D003497:Cyclic N-Oxides; D016166:Free Radical Scavengers; D007093:Imidazoles; D008163:Luminescent Measurements / Q000379:methods* / Q000592:standards; D008165:Luminol / Q000031:analogs & derivatives*; D009566:Nitrates / Q000037:antagonists & inhibitors*; D030421:Peroxynitrous Acid; D011794:Quercetin; D013113:Spin Labels; D014443:Tyrosine
D000975:Antioxidants; D003497:Cyclic N-Oxides; D016166:Free Radical Scavengers; D007093:Imidazoles; D009566:Nitrates; D013113:Spin Labels; C081614:L 012; D030421:Peroxynitrous Acid; C079391:2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide; D000082:Acetaminophen; D014443:Tyrosine; D008165:Luminol; D011794:Quercetin; C001803:tempol