A peroxyoxalate chemiluminescence-based assay for the evaluation of hydrogen peroxide scavenging activity employing 9,10-diphenylanthracene as the fluorophore.
BACKGROUND
A simple, rapid, sensitive, and enzyme-free analytical method for estimating scavenging of hydrogen peroxide (H2O2) was developed.


METHODS
Peroxyoxalate chemiluminescence (POCL) was measured, using 9,10-diphenylanthracene as fluorophore.


RESULTS
The chemiluminescence signal was found to be linear in response to increasing amounts of H2O2 in ethyl acetate/acetonitrile (9:1) (r2 = .9990), within a range of concentrations varying from 9.0 to 72.0 microM. In contrast, acetonitrile was highly unsuitable because of poor linearity (r2 = .3736) and poor signal stability. The linearity of POCL inhibition, as a measure of H2O2 scavenging, was tested employing well-known, lipid-soluble antioxidants, including beta-carotene, butylated hydroxytoluene (BHT) and alpha-tocopherol, and also the more polar flavonol quercetin, and the water-soluble L-ascorbic acid (AA). Under the experimental conditions, the corresponding values of H2O2 scavenging activity (SA(HP)) for quercetin, beta-carotene, alpha-tocopherol, and L-AA were 19.1 +/- 0.4, 70.9 +/- 20.1, 8.4 +/- 0.4, and 44.8 +/- 5.6 x 10(-3) microM(-1)


CONCLUSIONS
The data establish the assay as a method for assessing the H2O2 quenching activity of lipid-soluble antioxidants.
Laboratory of Chemistry of Natural Products, Mediterranean Agronomic Institute of Chania (MAICh), P.O. Box 85, 73100 Chania, Greece.
Anis
A
Arnous
Christos
C
Petrakis
Dimitris P
DP
Makris
Panagiotis
P
Kefalas
D000873:Anthracenes; D000975:Antioxidants / Q000737:chemistry; D001205:Ascorbic Acid / Q000737:chemistry; D002084:Butylated Hydroxytoluene / Q000737:chemistry; D005456:Fluorescent Dyes; D016166:Free Radical Scavengers / Q000737:chemistry*; D006861:Hydrogen Peroxide / Q000737:chemistry*; D008163:Luminescent Measurements; D015394:Molecular Structure; D010070:Oxalates / Q000737:chemistry*; D016877:Oxidants / Q000737:chemistry*; D011794:Quercetin / Q000737:chemistry; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D024502:alpha-Tocopherol / Q000737:chemistry; D019207:beta Carotene / Q000737:chemistry
D000873:Anthracenes; D000975:Antioxidants; D005456:Fluorescent Dyes; D016166:Free Radical Scavengers; D010070:Oxalates; D016877:Oxidants; D019207:beta Carotene; D002084:Butylated Hydroxytoluene; C046961:9,10-diphenylanthracene; C044622:peroxyoxalate; D011794:Quercetin; D006861:Hydrogen Peroxide; D024502:alpha-Tocopherol; D001205:Ascorbic Acid