Fluorescence-based assay for reactive oxygen species: a protective role for creatinine.
Attack by reactive oxygen species leads to a decay in phycoerythrin fluorescence emission. This phenomenon provides a versatile new assay for small molecules and macromolecules that can function as protective compounds. With 1-2 x 10(-8) M phycoerythrin, under conditions where peroxyl radical generation is rate-limiting, the fluorescence decay follows apparent zero-order kinetics. On reaction with HO., generated with the ascorbate-Cu2+ system, the fluorescence decays with apparent first-order kinetics. Examination of the major components of human urine in this assay confirms that at physiological concentrations, urate protects against both types of oxygen radicals. A novel finding is that creatinine protects efficiently by a chelation mechanism against radical damage in the ascorbate-Cu2+ system at creatinine, ascorbate, and Cu2+ concentrations comparable to those in normal urine. Urate and creatinine provide complementary modes of protection against reactive oxygen species in the urinary tract.
Glazer
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Department of Microbiology and Immunology, University of California, Berkeley 94720.
D001205:Ascorbic Acid; D003300:Copper; D003404:Creatinine / Q000502:physiology*; D005609:Free Radicals; D006801:Humans; D007700:Kinetics; D010100:Oxygen / Q000378:metabolism*; D010799:Phycoerythrin / Q000032:analysis; D013050:Spectrometry, Fluorescence
D005609:Free Radicals; D010799:Phycoerythrin; D003300:Copper; D003404:Creatinine; D001205:Ascorbic Acid; D010100:Oxygen