Abstract
Several methods have been developed to assess the radical scavenging activity. Among them, the 2,2-diphenyl-1-picrylhydrazyl (DPPH) spectrophotometric method is one of the most widely applied and is appreciated for its reliability. In this study, a comparison of two spectroscopic methods (electron paramagnetic resonance (EPR) and ultraviolet–visible (UV–Vis) spectroscopy) was performed analysing the spectroscopic features of DPPH in mixed ethanol/water solution and the free radical scavenging properties of myrtle leaves extracts and citrus juices. When DPPH was dissolved in mixed solvents, EPR enabled to identify the aggregation phenomena that occur when high amounts of water were employed. On the contrary, UV–Vis revealed only small differences in the absorption maximum among solutions with increasing water contents, without detecting aggregation. EC50 values of myrtle leaf extracts and citrus juices calculated from UV–Vis data were lower than those calculated with EPR. In myrtle extracts, the DPPH depletion measured by UV–Vis was not concentration dependent, revealing the interference in the analysis of the decomposition products of the antioxidants, which absorb at 517 nm. EPR spectroscopy was proven to be most reliable with all types of matrix since it is not dependent on the chemical composition of the extract.
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References
Brand-Williams W, Cuvelier ME, Berset C (1995) Use of a free radical method to evaluate antioxidant activity. Food Sci Technol LWT 28(1):25–30
Di Stefano R, Cravero MC (1991) Metodi per lo studio dei polifenoli dell’uva. Riv Viticol Enol 44:37–45
Dos Santos AB, Siqueira Silva DH, Da Silva Bolzani V, Avila Santos L, Schmidt TM, Baffa O (2009) Antioxidant properties of plant extracts: an EPR and DFT comparative study of the reaction with DPPH, TEMPOL and spin trap DMPO. J Braz Chem Soc 20:1483–1492
Gardner PT, White TAC, McPhail DB, Duthie GC (2000) The relative contributions of vitamin C, carotenoids and phenolics to the antioxidant potential of fruit juices. Food Chem 68:471–474
Huang D, Ou B, Pior RL (2005) The chemistry behind antioxidant capacity assays. J Agric Food Chem 53:1841–1856
Ladaniya M (2008) Citrus fruit—biology, technology and evaluation. Academic Press, San Diego
Magalhães LM, Segundo MA, Reis S, Lima JFLC (2008) Methodological aspects about in vitro evaluation of antioxidant properties. Anal Chim Acta 613:1–19
Maisuthisakul P, Suttajit M, Pongsawatmanit R (2007) Assessment of phenolic content and free radical scavenging capacity of some Thai indigenous plants. Food Chem 100:1409–1418
Moon JK, Shibamoto T (2009) Antioxidant assays for plant and food components. J Agric Food Chem 57:1655–1666
Oszmianski J, Wolniak M, Wojdylo A, Wawer I (2008) Influence of apple purée preparation and storage on polyphenol contents and antioxidant activity. Food Chem 107(4):1473–1484
Ozcelik B, Lee JH, Min DB (2003) Effects of light, oxygen, and pH on the absorbance of 2,2-diphenyk-1-picrylhydrazyl. J Food Sci 68:487–490
Polovka M (2006) EPR spectroscopy: a tool to characterize stability and antioxidant properties of foods. J Food Nutr Res 45:1–11
Polovka M, Brezová V, Stasko A (2003) Antioxidant properties of tea investigated by EPR spectroscopy. Biophys Chem 106(1):39–56
Polovka M, Brezová V, Stasko A, Mazúr M, Suhaj M, Simko P (2006) EPR investigations of gamma-irradiated ground black pepper. Radiat Phys Chem 75(2):309–321
Rapisarda P, Tomaino A, Cascio RL, Bonina F, De Pasquale A, Saija A (1999) Antioxidant effectiveness as influenced by phenolic content of fresh orange juices. J Agric Food Chem 47:4718–4723
Romani A, Pinelli P, Malinucci N, Vincieri FF, Tattini M (1999) Identification and quantitation of polyphenols in leaves of Myrtus communis L. Chromatographia 49:17–20
Romani A, Coinu R, Carta S, Pinelli P, Galardi G, Vincieri FF, Franconi F (2004) Evaluation of antioxidant effect of different extracts of Myrtus communis L. Free Rad Res 38:97–103
Sanchez-Moreno C (2002) Methods used to evaluate the free radical scavenging activity in foods and biological systems. Food Sci Technol Int 8:121–137
Sharma OP, Bhat TK (2009) DPPH antioxidant assay revisited. Food Chem 113(4):1202–1205
Smirnoff N (1996) The function and metabolism of ascorbic acid in plants. Ann Bot 78:661–669
Stasko A, Brezova V, Biskupic S, Misik V (2007) The potential pitfalls of using 1,1-diphenyl-2-picrylhydrazyl to characterize antioxidants in mixed water solvents. Free Rad Res 41:379–390
Tuberoso CIG, Rosa A, Bifulco E, Melis MP, Atzeri A, Pirisi FM, Dessì MA (2010) Chemical composition and antioxidant activities of Myrtus communis L. berries extracts. Food Chem 123 (4):1242–1251
Tzika ED, Papadimitriou V, Sotiroudis TG, Xenakis A (2008) Antioxidant properties of fruits and vegetables shots and juices: an electron pramagnetic resonace study. Food Biophys 3(1):48–53
Zafra-Stone S, Yasmin T, Bagchi M, Chatterjee A, Vinson JA, Bagch D (2007) Berry anthocyanins as novel antioxidants in human health and disease prevention. Mol Nutr Food Res 51:675–683
Acknowledgments
This work was partially supported by the “MIUR/CNR-Progetto Agroalimentare, Ambiente e Salute.” The authors would like to thank Mr. Giovanni Ligios for the technical assistance.
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Sanna, D., Delogu, G., Mulas, M. et al. Determination of Free Radical Scavenging Activity of Plant Extracts Through DPPH Assay: An EPR and UV–Vis Study. Food Anal. Methods 5, 759–766 (2012). https://doi.org/10.1007/s12161-011-9306-1
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DOI: https://doi.org/10.1007/s12161-011-9306-1