Lucigenin and linoleate enhanced chemiluminescent assay for superoxide dismutase activity.
The xanthine/xanthine oxidase dependent chemiluminescence was enhanced both by lucigenin and linoleate to create a sensitive, specific, and rapid chemiluminescent method for superoxide dismutase (SOD) activity determination. A pH optimum at around 10.0 was found both for the chemiluminescence and its inhibition by SOD. At this pH, a linear inhibition response to concentrations from 0.01 to 100 ng/ml of bovine Cu,Zn SOD could be established, with a 50% inhibitory concentration of 0.75 ng/ml. As little as 0.17 fmol of Cu,Zn SOD per test can be detected. With a slightly lower sensitivity, the method is operative at pH 7.4, too. Both Cu,Zn SOD and Mn SOD can be assayed. The rationale of the assay is in combining a superoxide-producing enzymatic system with linoleate amplification to enhance the sensitivity of the chemiluminescence to inhibition by SOD activity. Applicability of the method to biological samples was tested with a standard addition experiment.
Department of Physiology, University of Turku, Finland.
J K
JK
Laihia
C T
CT
Jansén
M
M
Ahotupa
D000166:Acridines / Q000494:pharmacology*; D000818:Animals; D005609:Free Radicals; D005979:Glutathione Peroxidase / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D019787:Linoleic Acid; D008041:Linoleic Acids / Q000494:pharmacology*; D008163:Luminescent Measurements; D051381:Rats; D012643:Selenium / Q000494:pharmacology; D018038:Sodium Selenite; D013482:Superoxide Dismutase / Q000032:analysis*; D019820:Xanthine; D014969:Xanthine Oxidase / Q000378:metabolism; D014970:Xanthines / Q000378:metabolism
D000166:Acridines; D005609:Free Radicals; D008041:Linoleic Acids; D014970:Xanthines; D019820:Xanthine; C033472:10,10'-dimethyl-9,9'-biacridinium; D019787:Linoleic Acid; D005979:Glutathione Peroxidase; D013482:Superoxide Dismutase; D014969:Xanthine Oxidase; D012643:Selenium; D018038:Sodium Selenite