Nitroxidation, nitration, and oxidation of a BODIPY fluorophore by RNOS and ROS

• Published in: Nitric oxide 2006-09, Vol.15 (2), p.163-176
• Main Authors: Nicolescu, Adrian C., Li, Qian, Brown, Laurie, Thatcher, Gregory R.J.
• Format: Article
• Language: English
• Subjects: Antioxidant; Boron Compounds - chemistry; Fluorescent Dyes - chemistry; Fluorescent probe; Free Radical Scavengers - chemistry; Molecular Probes - chemistry; Nitration; Nitrogen - chemistry; Nitroxidation; Oxidation; Oxidation-Reduction; Oxygen - chemistry; Peroxynitrite; Peroxynitrous Acid - chemistry; Reactive Nitrogen Species - chemistry; Reactive Oxygen Species - chemistry; RNOS; ROS; Spectrometry, Fluorescence
• ISSN: 1089-8603; 1089-8611
• DOI: 10.1016/j.niox.2006.01.010

BODIPY C 11 581/591 (BODIPY 11) represents a sensitive probe for quantification of relative antioxidant capacity. However, the mechanism of BODIPY 11 fluorescence decay in the presence of reactive oxygen species (ROS) and reactive nitrogen oxide species (RNOS) requires clarification. Azo-initiators provide a continuous source of peroxyl radicals that in simple, aerobic, homogeneous, buffered solution simulate lipid peroxyl radical formation. Inhibition of BODIPY 11 fluorescence decay was assayed and quantified for several families of antioxidants, including phenols, NO donors, and thiols. Fluorescence decay of BODIPY 11 in these systems demonstrated similar patterns of antioxidant activity to those observed in classical oxygen pressure measurements, and provided a readily applied quantification of antioxidant capacity and mechanistic information, which was analyzed by measurement of induction periods, initial rates, and net oxidation. LC/MS analysis confirmed that peroxyl radical-induced irreversible fluorescence decay of the BODIPY 11 fluorophore is due to oxidative cleavage of the activated phenyldiene side chain. The behavior of BODIPY 11 towards RNOS was more complex, even in these simple systems. Incubation of BODIPY 11 with bolus peroxynitrite or a sydnonimine peroxynitrite source produced a variety of novel products, characterized by LC/MS, derived from oxidative cleavage, nitroxidation, and nitration reactions. The “NO scavenger” PTIO reinforced the antioxidant activity of NO, and inhibited BODIPY 11 oxidation induced by the sydnonimine. These observations suggest that BODIPY 11 is a well-behaved fluorescence probe for peroxidation and antioxidant studies, but that for study of RNOS even co-application of fluorescence decay with LC/MS measurements requires careful analysis and interpretation.