Superoxide anion formation from lucigenin: an electron spin resonance spin-trapping study.
Lucigenin (LC2+) is frequently used as a superoxide probe. To detect superoxide, lucigenin must be reduced to the lucigenin cation radical (LC.+). We show, using the phosphorylated spin trap 5-diethoxyphosphoryl-5-methyl-1-pyrroline N-oxide (DEPMPO), that lucigenin stimulates NADPH-dependent superoxide production by endothelial nitric oxide synthase (eNOS). The formation of the DEPMPO-superoxide adduct is calcium/calmodulin independent. DEPMPO-superoxide adduct formation is inhibited by diphenyleneiodonium and is abolished by superoxide dismutase. It is likely that eNOS/NADPH can reduce lucigenin to LC.+ which reduces oxygen to superoxide. Consequently, lucigenin cannot be used to measure superoxide formation.
Vásquez-Vivar
J
J
Biophysics Research Institute, Medical College of Wisconsin, Milwaukee 53226-0509, USA.
Hogg
N
N
Pritchard
K A
KA
Martasek
P
P
Kalyanaraman
B
B
D000166:Acridines / Q000737:chemistry / Q000378:metabolism*; D000818:Animals; D002417:Cattle; D004578:Electron Spin Resonance Spectroscopy; D015335:Molecular Probes; D009249:NADP / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000302:isolation & purification / Q000378:metabolism; D010084:Oxidation-Reduction; D013481:Superoxides / Q000737:chemistry / Q000378:metabolism*
D000166:Acridines; D015335:Molecular Probes; D013481:Superoxides; C033472:10,10'-dimethyl-9,9'-biacridinium; D009249:NADP; D019001:Nitric Oxide Synthase