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 × 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, A. N. Fluorescence‐based assay for reactive oxygen species: a protective role for creatinine. FASEB J. 2: 2487‐2491; 1988.