Inhibition by L-ascorbic acid and other antioxidants of the 2.2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) oxidation catalyzed by peroxidase: a new approach for determining total antioxidant status of foods.
The accumulation of 2.2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) radical catalyzed by peroxidase can be inhibited by the presence of L-ascorbic acid in the reaction medium, this inhibition delaying the accumulation of the ABTS radical and giving rise to a lag time. A kinetic approach to explain this lag time is presented, which also makes it possible to determine the amount of L-ascorbic acid in the reaction medium. The stoichiometry of the system was determined as 1 mol of L-ascorbic reducing 2 mol of ABTS radicals. L-Ascorbic acid is not the only compound to have this ability, since other antioxidant compounds also react with the ABTS radical. We studied the ABTS/H2O2/horseradish peroxidase system in the presence of L-ascorbic acid and other antioxidant compounds. The influence of such factors as pH, enzyme concentration, and L-ascorbic acid concentration was studied. A good correlation between the lag time and the L-ascorbic acid present in the medium was observed, and under optimal conditions, the method could determine as little as 0.65 nmol of L-ascorbic acid. Based on our findings, we propose a method to measure the total antioxidant activity of different compounds related to L-ascorbic acid and apply this method to determining the total antioxidant activity present in fruit juices.
Arnao
M B
MB
Departamento de Biología Vegetal, Facultad de Biología, Universidad de Murcia, Spain.
Cano
A
A
Hernández-Ruiz
J
J
García-Cánovas
F
F
Acosta
M
M
D000975:Antioxidants / Q000494:pharmacology*; D001205:Ascorbic Acid / Q000494:pharmacology*; D052160:Benzothiazoles; D002138:Calibration; D002384:Catalysis; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005504:Food Analysis / Q000379:methods*; D005609:Free Radicals; D006735:Horseradish Peroxidase / Q000037:antagonists & inhibitors*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008786:Metmyoglobin; D010084:Oxidation-Reduction; D013053:Spectrophotometry; D013451:Sulfonic Acids / Q000737:chemistry*
D000975:Antioxidants; D052160:Benzothiazoles; D004791:Enzyme Inhibitors; D005609:Free Radicals; D013451:Sulfonic Acids; D008786:Metmyoglobin; C002502:2,2'-azino-di-(3-ethylbenzothiazoline)-6-sulfonic acid; D006735:Horseradish Peroxidase; D001205:Ascorbic Acid