Assay dilution factors confound measures of total antioxidant capacity in polyphenol-rich juices.

UNLABELLED: The extent to which sample dilution factor (DF) affects total antioxidant capacity (TAC) values is poorly understood. Thus, we examined the impact of DF on the ORAC, FRAP, DPPH, and total phenols (TP) assays using pomegranate juice (PJ), grape juice (GJ), selected flavonoids, ascorbic acid, and ellagic acid. For ORAC, GJ was comparable to PJ at DF 750, but at DF 2000, the ORAC value of GJ was 40% more than PJ. Increasing DF increased GJ and PJ, DPPH, TP, and FRAP values 11% and 14%, respectively. Increased test concentrations of quercetin and catechin resulted in 51% and 126% greater ORAC values, but decreased naringenin by 68%. Flavonoids, but not ellagic acid or ascorbic acid, may contribute to the dilution effect on the variation of final TAC values. Thus, reporting TAC or TP using a single DF may introduce uncertainty about the confidence of TAC assay values, especially when comparing different juices. These results underscore the importance of using compatible test standards for reporting TAC values. PRACTICAL APPLICATION: Total antioxidant capacity (TAC) values such as the ORAC assay are increasingly used for comparison of polyphenol-rich foods and beverages. Choice of standards and test concentrations, even within the linear range of standards, may introduce variation probably due to synergy/antagonism between antioxidant and thereby, confound final TAC values. Thus, test concentration or dilution factors of samples should be considered in the design of TAC assays and interpretation of their results.