Action of DCFH and BODIPY as a probe for radical oxidation in hydrophilic and lipophilic domain.
Fluorogenic probes such as 2',7'-dichlorofluorescin (DCFH) have been extensively used to detect oxidative events and to measure antioxidant capacity. At the same time, however, the inherent drawbacks of these probes such as non-specificity towards oxidizing species have been pointed out. The present study was carried out to analyze the action and dynamics of 4, 4-difluoro-5-(4-phenyl-1,3-butadienyl)-4-bora-3a,4a-diaza-s-indacene-3-undecanoic acid (BODIPY) and DCFH as a fluorescent probe in the free radical-mediated lipid peroxidation in homogeneous solution, aqueous suspensions of liposomal membranes and LDL and plasma. The rate constant for the reaction of BODIPY with peroxyl radicals was estimated as 6.0 x 10(3) M(-1) s(-1), which makes BODIPY kinetically an inefficient probe especially in the presence of potent radical-scavenging antioxidants such as tocopherols, but a convenient probe for lipid peroxidation. On the other hand, the reactivity of DCFH toward peroxyl radicals was as high as Trolox, a water-soluble analogue of alpha-tocopherol. Thus, DCFH is kinetically more favored probe than BODIPY and could scavenge the radicals within lipophilic domain as well as in aqueous phase. The partition coefficients for BODIPY and DCFH were obtained as 4.57 and 2.62, respectively. These results suggest that BODIPY may be used as an efficient probe for the free radical-mediated oxidation taking place in the lipophilic domain, especially after depletion of alpha-tocopherol, while it may not be an efficient probe for detection of aqueous radicals.
Yoshida
Yasukazu
Y
Human Stress Signal Research Center (HSSRC), National Institute of Advanced Industrial Science and Technology (AIST), Ikeda, Osaka 563-8577, Japan. yoshida-ya@aist.go.jp
Shimakawa
Shuichi
S
Itoh
Nanako
N
Niki
Etsuo
E
D000975:Antioxidants / Q000737:chemistry; D001896:Boron Compounds / Q000737:chemistry / Q000494:pharmacology*; D002462:Cell Membrane / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004578:Electron Spin Resonance Spectroscopy; D005452:Fluoresceins / Q000737:chemistry / Q000494:pharmacology*; D005456:Fluorescent Dyes / Q000494:pharmacology; D005609:Free Radicals; D006801:Humans; D007700:Kinetics; D015227:Lipid Peroxidation; D008077:Lipoproteins, LDL / Q000378:metabolism; D008081:Liposomes / Q000378:metabolism; D008956:Models, Chemical; D010100:Oxygen / Q000378:metabolism; D017434:Protein Structure, Tertiary; D013050:Spectrometry, Fluorescence; D013997:Time Factors
C095489:4,4-difluoro-4-bora-3a,4a-diaza-s-indacene; D000975:Antioxidants; D001896:Boron Compounds; D005452:Fluoresceins; D005456:Fluorescent Dyes; D005609:Free Radicals; D008077:Lipoproteins, LDL; D008081:Liposomes; C029569:diacetyldichlorofluorescein; D010100:Oxygen