ABSTRACT
A fluorescent probe, DPPEC (1,2-dipalmitoylglycerophosphorylethanolamine labeled with coumarin) was developed for detecting hydroxyl radical (*OH) in lipid membranes. The coumarin moiety contributes to the fluorescent detection of *OH and the phospholipids moiety gives a driving force to localize the probe in lipid membranes. DPPEC in liposomal membranes rapidly reacted with *OH and increased the fluorescence intensity, depending on the concentration of *OH. The increase in the fluorescence intensity induced by *OH was effectively suppressed by the addition of DMSO. The probe exhibited a higher fluorescence response to *OH over other reactive oxygen species, such as hydrogen peroxide, nitric oxide, peroxynitrite, alkylperoxyl radical, and hypochlorite. DPPEC would be useful as a new type of fluorescent probe that can localize in lipid membranes and detect *OH efficiently.
Subject(s)
Coumarins/chemistry , Ethanolamines/chemistry , Fluorescent Dyes/chemistry , Glycerol/analogs & derivatives , Hydroxyl Radical/analysis , Liposomes/chemistry , Membranes, Artificial , Phospholipids/chemistry , Ethanolamines/chemical synthesis , Glycerol/chemical synthesis , Glycerol/chemistry , Molecular Structure , Phospholipids/chemical synthesis , Reproducibility of Results , Time FactorsABSTRACT
A novel fluorescent probe, 7-hydroxy-2-oxo-N-(2-(diphenylphosphino)ethyl)-2H-chromene-3-carboxamide (DPPEA-HC) was developed for use in monitoring hydrogen peroxide (H2O2) production. DPPEA-HC, which consists of a diphenylphosphine moiety and a 7-hydroxycoumarin moiety, reacts with H2O2 to form DPPEA-HC oxide, which is analogous to the reaction of triphenylphosphine with hydroperoxides such as H2O2 to form triphenylphosphine oxide. Photoinduced electron transfer (PET) was applied in the design of DPPEA-HC. Since the diphenylphosphine moiety and the 7-hydroxycoumarin moiety would act as the PET donor and the acceptor, respectively, it would be expected that DPPEA-HC would rationally cancel the PET process via the formation of DPPEA-HC oxide, based on the calculated energy levels of the donor and the acceptor moieties using the B3LYP/6-31G*//AM1 method. The fluorescence intensity of DPPEA-HC increased on the addition of a H2O2 solution in 100 mM sodium phosphate buffer (pH7.4), as predicted from the energy level calculation and a good correlation between increase in the fluorescence of DPPEA-HC and the concentration of H2O2 was observed. DPPEA-HC was also fluoresced by H2O2, which was enzymatically produced in xanthine/xanthine oxidase/superoxide dismutase (XA/XOD/SOD) system. The increase in the fluorescence of DPPEA-HC in the presence of H2O2 immediately ceased on the addition of catalase (CAT), which catalyzes the disproportionation of H2O2. In addition, DPPEA-HC was found to have a much higher selectivity for H2O2 and a greater resistance to autoxidation than 2',7'-dichlorodihydrofluoresein (DCFH). Time-resolved fluorescence measurements of DPPEA-HC and DPPEA-HC oxide confirmed that the fluorescence off/on switching mechanism of DPPEA-HC is based on the PET on/off control.
Subject(s)
Benzopyrans/chemistry , Electron Transport , Fluorescent Dyes/chemistry , Hydrogen Peroxide/analysis , Hydrogen Bonding , Models, MolecularABSTRACT
A novel fluorescent probe, the detection mechanism of which is based on the 'on-off' switching of a FRET triggered by the *OH-induced cleavage of a DNA strand, has been developed for the ratiometric imaging of *OH.
