SPECTROPHOTOMETRIC DETERMINATION OF ANTIOXIDANT ACTIVITY

The spectrophotometric technique for total antioxidant activity (TAA)( 1,2) measures the relative abilities of antioxidants to scavenge the 2 ,2'-azino-bis(3-ethylbenz-thiazoline-6-sulfonic acid) (ABTS) radical c ation (ABTS(.+)) in comparison with the antioxidant potency of standar d amounts of Trolox, the water-soluble vitamin E analogue. This method is based on the progressive consumption of antioxidant activity by AB TS(.+) as it is generated in the reaction cuvette and can be automated with a spectrophotometric analyzer. Several different analytical stra tegies are possible using the same reagents, enabling the assay system to be used to determine the antioxidant activity of plasma, saliva, l ipoprotein fractions, foods and beverages. To determine the activity o f pure antioxidant substances, a hydrogen peroxide concentration of 75 mu M is used, together with a 6 min measuring time. For biological sa mples with endogenous peroxidase activity the hydrogen peroxide concen tration is increased fivefold and the measuring time shortened to 3.25 min. Assays with improved sensitivity are described for low-density l ipoprotein (LDL) preparations and saliva. Use of a spectrophotometric endpoint makes the assay simple to carry out without special laborator y equipment. Measurement at 734 nm avoids a range of potential interfe ring factors, such as sample turbidity and non-specific absorbance by sample constituents. Current applications of the ABTS antioxidant assa y are described and discussed.