title: Insights on antioxidant assays for biological samples based on the reduction of copper complexes-the importance of analytical conditions.
abstract: Total antioxidant capacity assays are recognized as instrumental to establish antioxidant status of biological samples, however the varying experimental conditions result in conclusions that may not be transposable to other settings. After selection of the complexing agent, reagent addition order, buffer type and concentration, copper reducing assays were adapted to a high-throughput scheme and validated using model biological antioxidant compounds of ascorbic acid, Trolox (a soluble analogue of vitamin E), uric acid and glutathione. A critical comparison was made based on real samples including NIST-909c human serum certified sample, and five study samples. The validated method provided linear range up to 100 µM Trolox, (limit of detection 2.3 µM; limit of quantification 7.7 µM) with recovery results above 85% and precision <5%. The validated developed method with an increased sensitivity is a sound choice for assessment of TAC in serum samples.
authors: REQUIMTE, Departamento de Química, Faculdade de Farmácia, Universidade do Porto, Rua Jorge Viterbo Ferreira, 228, 4050-313 Porto, Portugal. micf09213@ff.up.pt.
authors: Sara S
authors: SS
authors: Marques
authors: REQUIMTE, Departamento de Química, Faculdade de Farmácia, Universidade do Porto, Rua Jorge Viterbo Ferreira, 228, 4050-313 Porto, Portugal. luismagalhaes@ff.up.pt.
authors: Luís M
authors: LM
authors: Magalhães
authors: REQUIMTE, Departamento de Química, Faculdade de Farmácia, Universidade do Porto, Rua Jorge Viterbo Ferreira, 228, 4050-313 Porto, Portugal. ivtoth@ff.up.pt.
authors: Ildikó V
authors: IV
authors: Tóth
authors: REQUIMTE, Departamento de Química, Faculdade de Farmácia, Universidade do Porto, Rua Jorge Viterbo Ferreira, 228, 4050-313 Porto, Portugal. msegundo@ff.up.pt.
authors: Marcela A
authors: MA
authors: Segundo
mesh_terms: D000975:Antioxidants / Q000032:analysis*; D001205:Ascorbic Acid / Q000097:blood; D001774:Blood Chemical Analysis / Q000379:methods*; D002839:Chromans / Q000097:blood; D003300:Copper / Q000737:chemistry*; D005978:Glutathione / Q000097:blood; D057166:High-Throughput Screening Assays / Q000379:methods; D006801:Humans; D010084:Oxidation-Reduction; D014527:Uric Acid / Q000097:blood
chemical_list: D000975:Antioxidants; D002839:Chromans; D014527:Uric Acid; D003300:Copper; D005978:Glutathione; D001205:Ascorbic Acid; C010643:6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid