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ARS Home » Northeast Area » Boston, Massachusetts » Jean Mayer Human Nutrition Research Center On Aging » Research » Publications at this Location » Publication #193177
Title: TOTAL ANTIOXIDANT PERFORMANCE (TAP): A VALIDATED FLUORESCENCE ASSAY FOR THE MEASUREMENT OF PLASMA OXIDIZABILITY

Author
item BERETTA, GIANGIACOMO - UNIVERSITY OF MILAN
item ALDINI, GIANCARLO - UNIVERSITY OF MILAN
item FACINO, ROVERTO - UNIVERSITY OF MILAN
item Russell, Robert
item KRINSKY, NORMAN - TUFTS/HNRCA
item YEUM, KYUNG-JIN - TUFTS/HNRCA

Submitted to: Analytical Chemistry
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/1/2006
Publication Date: 7/1/2006
Citation: Beretta, G., Aldini, G., Facino, R.M., Russell, R., Krinsky, N.I., Yeum, K. 2006. Total antioxidant performance (tap): a validated fluorescence assay for the measurement of plasma oxidizability. Analytical Chemistry. 354(2):290-8.

Interpretive Summary: The antioxidant activity of human blood was determined by following the oxidation of the lipid soluble oxidation indicating dye. Damage was caused by a substance producing damage in the fat soluble portion of blood. The results are expressed as a Total Antioxidant Performance (TAP) value, based on the protection of oxidation of the lipid soluble indicating dye. The fittingness of the test was evaluated on the basis of its accuracy, producing same results repeatedly, and identifying exactly. The differences of the test during days and within day were both under 5%. The addition of lipid did not show any meaningful change in the TAP value. Also, the lipid soluble oxidation indicating dye was not damaged by light. The TAP values of 6 blood samples from healthy people were measured and determined for the relationship with the blood water- and fat-soluble antioxidants (uric acid and vitamin C, vitamin E, carotenoids). Meaningful relationships were found between TAP and uric acid and vitamin E. The results show that the TAP test is good to measure the antioxidant activity of blood antioxidants present in both the fat-soluble and water-soluble portions.

Technical Abstract: The antioxidant capacity of human plasma was determined by following the oxidation kinetics of the lipid soluble fluorescent marker, BODIPY using MeO-AMVN as the lipophilic radical initiator. The results are expressed as a Total Antioxidant Performance (TAP) value, based on the inhibition of BODIPY oxidation, as determined by the appearance of green fluorescence, with respect to a control sample (phosphatidylcholine with or without delipidized human serum). The suitability of the assay was evaluated on the basis of its precision, reproducibility and specificity. The intra- and inter-assays CV% were both under 5%. The addition of a representative substrate of plasma peroxidation, phosphatidylcholine up to 750 mcg/ml did not induce significant changes in the TAP value. Also, BODIPY photo-oxidation was not observed during the experimental time-course (220 min). The TAP values of 6 plasma samples from healthy donors were measured and correlated with the main plasma water- and lipid-soluble antioxidants (uric acid and ascorbic acid, a-tocopherol, carotenoids) and lipid profiles. Significant correlations were found between TAP and uric acid (R=0.97, p<0.05) and cholesterol-adjusted a-tocopherol (R=0.93, p<0.01). The results confirm that the TAP assay is suitable to measure the antioxidant activity of plasma antioxidants localized in both the lipophilic and hydrophilic compartments.