title: The intracellular iron sensor calcein is catalytically oxidatively degraded by iron(II) in a hydrogen peroxide-dependent reaction.
abstract: The fluorescent metal chelating dye calcein is used to obtain an estimate of cellular iron levels and to measure the kinetics of the entry of chelators and chelating drugs into cells. Under reducing conditions in the presence of ascorbic acid, such as that would be present in the cell, the Fe(II)-calcein complex was rapidly formed with a rate constant of 3 x 10(5) M(-1) s(-1). A slower iron-dependent catalytic degradation of calcein also occurred that resulted in the formation of a non-fluorescent calcein product. The Fe(II)-catalyzed degradation of calcein was largely, but not completely, prevented by catalase. Electron paramagnetic resonance spin trapping experiments showed that the Fe(II)-calcein complex promoted formation of hydroxyl or a hydroxyl radical-like species. Together these results indicated that Fe(II) catalyzed the degradation of calcein through both hydrogen peroxide, and to a lesser extent, non-hydrogen peroxide-dependent pathways. The iron-calcein complexes that were responsible for the degradation of calcein were likely high valence oxidizing iron-oxo species such as perferryl or ferryl complexes that were redox cycled by ascorbic acid. Thus, the use of calcein as an intracellular iron-sensing indicator may yield misleading results due to its degradation under certain conditions.
authors.affiliation: Faculty of Pharmacy, University of Manitoba, Winnipeg, Manitoba, Canada R3T 2N2. b_hasinoff@umanitoba.ca
authors.forename: Brian B
authors.initials: BB
authors.lastname: Hasinoff
mesh_terms: D001205:Ascorbic Acid / Q000737:chemistry; D002374:Catalase / Q000378:metabolism / Q000494:pharmacology; D002384:Catalysis; D002614:Chelating Agents / Q000737:chemistry / Q000378:metabolism; D004578:Electron Spin Resonance Spectroscopy; D005296:Ferrous Compounds / Q000737:chemistry* / Q000378:metabolism; D005452:Fluoresceins / Q000737:chemistry* / Q000378:metabolism; D005456:Fluorescent Dyes / Q000737:chemistry* / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000737:chemistry*; D017665:Hydroxyl Radical / Q000737:chemistry; D007501:Iron / Q000032:analysis / Q000737:chemistry*; D007700:Kinetics; D010084:Oxidation-Reduction; D013050:Spectrometry, Fluorescence / Q000379:methods
chemical_list: D002614:Chelating Agents; D005296:Ferrous Compounds; D005452:Fluoresceins; D005456:Fluorescent Dyes; D017665:Hydroxyl Radical; D006861:Hydrogen Peroxide; D007501:Iron; D002374:Catalase; D001205:Ascorbic Acid; C007740:fluorexon