title: Use of rapid-scan EPR to improve detection sensitivity for spin-trapped radicals.
abstract: The short lifetime of superoxide and the low rates of formation expected in vivo make detection by standard continuous wave (CW) electron paramagnetic resonance (EPR) challenging. The new rapid-scan EPR method offers improved sensitivity for these types of samples. In rapid-scan EPR, the magnetic field is scanned through resonance in a time that is short relative to electron spin relaxation times, and data are processed to obtain the absorption spectrum. To validate the application of rapid-scan EPR to spin trapping, superoxide was generated by the reaction of xanthine oxidase and hypoxanthine with rates of 0.1-6.0 μM/min and trapped with 5-tert-butoxycarbonyl-5-methyl-1-pyrroline-N-oxide (BMPO). Spin trapping with BMPO to form the BMPO-OOH adduct converts the very short-lived superoxide radical into a more stable spin adduct. There is good agreement between the hyperfine splitting parameters obtained for BMPO-OOH by CW and rapid-scan EPR. For the same signal acquisition time, the signal/noise ratio is >40 times higher for rapid-scan than for CW EPR. Rapid-scan EPR can detect superoxide produced by Enterococcus faecalis at rates that are too low for detection by CW EPR.
authors.affiliation: Department of Chemistry and Biochemistry, University of Denver, Denver, Colorado, USA.
authors.forename: Deborah G
authors.initials: DG
authors.lastname: Mitchell
authors.forename: Gerald M
authors.initials: GM
authors.lastname: Rosen
authors.forename: Mark
authors.initials: M
authors.lastname: Tseitlin
authors.forename: Breanna
authors.initials: B
authors.lastname: Symmes
authors.forename: Sandra S
authors.initials: SS
authors.lastname: Eaton
authors.forename: Gareth R
authors.initials: GR
authors.lastname: Eaton
mesh_terms: D004578:Electron Spin Resonance Spectroscopy / Q000379:methods; D013293:Enterococcus faecalis / Q000737:chemistry; D019271:Hypoxanthine / Q000737:chemistry; D057230:Limit of Detection; D011126:Polypropylenes / Q000737:chemistry; D019097:Spin Trapping / Q000379:methods*; D013481:Superoxides / Q000032:analysis* / Q000737:chemistry; D014969:Xanthine Oxidase / Q000737:chemistry
chemical_list: D011126:Polypropylenes; C545693:bis((3-methyldimethoxysilyl)propyl) polypropylene oxide; D013481:Superoxides; D019271:Hypoxanthine; D014969:Xanthine Oxidase