Skip to main content
SpringerLink
Log in

Measuring Antioxidant Capacity Using the ORAC and TOSC Assays

  • Protocol
  • First Online:
Advanced Protocols in Oxidative Stress II

Part of the book series: Methods in Molecular Biology ((MIMB,volume 594))

Abstract

Recent epidemiological studies have shown that there may be a link between oxidative stress and the development of several types of chronic diseases. Studies have also shown that diets rich in fruits and vegetables may decrease the incidence of cancer and other chronic diseases. The antioxidant activity of the phytochemicals these foods contain may be partially responsible for the decreased incidence of these diseases in people who regularly consume them. While there are several assays currently used to assess the antioxidant activity of phytochemicals and other antioxidant compounds, two are reviewed here in detail. The first is the oxygen radical absorbance capacity (ORAC) assay, which measures the decrease in fluorescence decay caused by antioxidants, and the second is the total oxyradical scavenging capacity (TOSC) assay, which measures the decrease in ethylene gas production caused by the inhibition of the thermal hydrolysis of ABAP (2,2′-Azobis(2-methyl-(propionamidine) dihydrochloride) by KMBA (α-keto-γ-(methylthio)butyric acid sodium salt) in the presence of antioxidant compounds. These two assays are discussed here, with an in depth review of their methodology and correlation.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 159.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Similar content being viewed by others

References

  1. Oxygen Radical Absorbance Capacity (ORAC) of Selected Foods (2007); Nutrient Data Laboratory, Agricultural Research Service, US Department of Agriculture, United States Department of Agriculture.

    Google Scholar 

  2. Ames BN, Gold LS, Willet WC (1995) The Causes and Prevention of cancer. Proc Natl Acad Sci USA 92:5258–5265

    Article  PubMed  CAS  Google Scholar 

  3. Ames BN, Shigenaga MK, Hagen TM (1993) Oxidants, antioxidants, and the degenerative diseases of aging. Proc Natl Acad Sci USA 90:7915–7922

    Article  PubMed  CAS  Google Scholar 

  4. Christen Y (2000) Oxidative stress and Alzheimer’s disease. Am J Clin Nutr 71:621S–629S

    PubMed  CAS  Google Scholar 

  5. Diaz MN, Frei B, Keaney JF Jr (1997) Antioxidants and atherosclerotic heart disease. N Eng J Med 337:408–416

    Article  CAS  Google Scholar 

  6. Lang AE, Lozano AM (1998) Parkinson’s disease. First of two parts. N Eng J Med 339:111–114

    Google Scholar 

  7. Packer L, Colman C (1999) The anti-oxidant miracle. Wiley, New York

    Google Scholar 

  8. Cao G, Prior RL (1998) Comparison of different analytical methods for assessing total antioxidant capacity of human serum. Clin Chem 44(6):1309–1315

    PubMed  CAS  Google Scholar 

  9. Halliwell B, Gutteridge JMC, Cross CE (1992) Free Radicals, antioxidants, and human disease: where are we now? J Clin Lab Med 119:598–620

    CAS  Google Scholar 

  10. Tomer DP (2003) Measuring parameters that are important in cancer prevention and treatment: assessing antioxidant activity and angiogenesis. Brigham Young University, Master’s Thesis

    Google Scholar 

  11. McLeman LD (2004) Evidence of synergistic intracellular antioxidant networking and antioxidant regulation of cellular lipid peroxidation in HL-60 acute promyelogenous leukemia. Master’s Thesis. Brigham Young University. 1-37.

    Google Scholar 

  12. O’Neill K, Murray B (2002) Power plants. Woodland Publishing, Utah

    Google Scholar 

  13. Cao G, Booth SL, Sadowski JA, Prior RL (1998) Increases in human plasma antioxidant capacity after consumption of controlled diets high in fruit and vegetables. Am J Clin Nutr 68:1081–1087

    PubMed  CAS  Google Scholar 

  14. Huang D, Ou B, Prior RL (2005) The chemistry behind antioxidant capacity assays. J Agric Food Chem 53:1841–1856

    Article  PubMed  CAS  Google Scholar 

  15. Ou B, Hampsch-Woodill M, Prior RL (2001) Development and validation of an improved oxygen radical absorbance capacity assay using fluorescein as the fluorescent probe. J Agric Food Chem 49(10):4619–4626

    Article  PubMed  CAS  Google Scholar 

  16. Cao G, Alessio H, Cutler R (1993) Oxygen-radical absorbance capacity assay for antioxidants. Free Radic Biol Med 14(3):303–311

    Article  PubMed  CAS  Google Scholar 

  17. Cao G, Sofic E, Prior RL (1996) Antioxidant capacity of tea and common vegetables. J Agric Food Chem 44:3426–3431

    Article  CAS  Google Scholar 

  18. Huang D, Ou B, Hampsch-Woodill M, Flanagan J, Prior RL (2002) High-throughput assay of oxygen radical absorbance capacity (orac) using a multichannel liquid handling system coupled with a microplate fluorescence reader in 96-well format. J Agric Food Chem 50:4437–4444

    Article  PubMed  CAS  Google Scholar 

  19. Ou B, Huang D, Hampsch-Woodill M, Flanagan JA, Deemer EK (2002) Analysis of antioxidant of common vegetables employing oxygen radical absorbance capacity (ORAC) and ferric reducing antioxidant power (FRAP) assays: a comparative study. J Agric Food Chem 50:3122–3128

    Article  PubMed  CAS  Google Scholar 

  20. Glazer AN (1990) Phycoerythrin fluorescence based assay for reactive oxygen species. Methods Enzymol 186:161–168

    Article  PubMed  CAS  Google Scholar 

  21. Cao G, Prior RL (1999) Measurement of oxygen radical absorbance capacity in biological samples. Methods Enzymol 299:50–62

    Article  PubMed  CAS  Google Scholar 

  22. Pellegrini N, Del Rio D, Colombi B, Bianchi M, Brighenti F (2003) Application of the 2, 2’-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) radical cation assay to a flow injection system for the evaluation of antioxidant activity of some pure compounds and beverages. J Agric Food Chem 51:260–264

    Article  PubMed  CAS  Google Scholar 

  23. Pulido R, Bravo L, Suara-Calixto F (2000) Antioxidant activity of dietary polyphenols as determined by a modified ferric reducing/antioxidant power assay. J Agric Food Chem 48:3396–3402

    Article  PubMed  CAS  Google Scholar 

  24. Aruoma OI (2003) Methodological considerations for characterizing potential antioxidant actions of bioactive components in plant foods. Mutat Res 523–524:9–20

    PubMed  Google Scholar 

  25. Gorinstein S, Martin-Belloso O, Katrich E, Lojek A, Ciz M, Gligelmo-Miguel N, Haruenkit R, Park YS, Jung ST, Trakhtenberg S (2003) Comparison of the contents of the main biochemical compounds and the antioxidant activity of some Spanish olive oils as determined by four different radical scavenging tests. J Nutr Biochem 14:154–159

    Article  PubMed  CAS  Google Scholar 

  26. Yokozawa T, Cho EJ, Hara Y, Kitani K (2000) Antioxidative activity of green tea treated with radical initiator 2, 2’-azobis(2-amidinopropane) dihydrochloride. J Agric Food Chem. 48:5068–5073

    Article  PubMed  CAS  Google Scholar 

  27. Prior RL, Hoang H, Gu L, Wu X, Bacchiocca M, Howard L, Hampsch-Woodill M, Huang D, Ou B, Jacob R (2003) Assays for hydrophilic and lipophilic antioxidant capacity (oxygen radical absorbance capacity (ORAC(FL))) of plasma and other biological and food samples. J Agric Food Chem 51:3273–3279

    Article  PubMed  CAS  Google Scholar 

  28. Dugas AJ Jr, Castaneda-Acosta J, Bonin GC, Price KL, Fischer NH, Winston GW (2000) Evaluation of the total peroxyl radical-scavenging capacity of flavonoids: structure-activity relationships. J Nat Prod 63:327–331

    Article  PubMed  CAS  Google Scholar 

  29. Regoli F, Winston GW (1999) Quantification of total oxidant scavenging capacity of antioxidants for peroxynitrite, peroxyl radicals, and hydroxyl radicals. Toxicol Appl Pharmacol 156:96–105

    Article  PubMed  CAS  Google Scholar 

  30. Adapted from http://www.bmglabtech.com/application-notes/fluorescence-intensity/orac-148.cfm. Consulted 7/9/08.

  31. Somogyi A, Rosta K, Pusztai P, Tulassay Z, Nagy G (2007) Antioxidant Measurements. Physiol Meas 28:R41–R55

    Article  PubMed  Google Scholar 

  32. Tomer DP, McLeman LD, Ohmine S, Scherer PM, Murray BK, O’Neill KL (2007) Comparison of the total oxyradical scavenging capacity and oxygen radical absorbance capacity assays. J Med Food 10(2):337–344

    Article  PubMed  CAS  Google Scholar 

  33. Lichtenthaler R, Marx F (2005) Total oxidant scavenging capacities of common european fruit and vegetable juices. J Agric Food Chem 53:103–110

    Article  PubMed  Google Scholar 

  34. Lichtenthaler R, Rodrigues RB, Maia JGS, Papagiannopoulos M, Fabricius H, Marx F (2005) Total oxidant scavenging capacities of Euterpe oleracea Mart. (Acai) fruits. Int J Food Sci Nutr 56(1):53–64

    Google Scholar 

  35. Company R, Serafim A, Cosson R, Camus L, Shillito B, Fiala-Medioni A, Bebianno MJ (2006) The effect of cadmium on antioxidant responses and the susceptibility to oxidative stress in the hydrothermal vent mussel Bathymodiolus azoricus. Marine Biol 148:817–825

    Article  CAS  Google Scholar 

  36. Giessing AMB, Mayer LM (2004) Oxidative coupling during gut passage in marine deposit-feeding invertebrates. Limnol Oceanogr 49(3):716–726

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Microbiology and Molecular Biology, Brigham Young University, Provo, UT, USA

    Andrew R. Garrett, Byron K. Murray, Richard A. Robison & Kim L. O’Neill

Authors
  1. Andrew R. Garrett
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Byron K. Murray
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Richard A. Robison
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Kim L. O’Neill
    View author publications

    You can also search for this author in PubMed Google Scholar

Editor information

Editors and Affiliations

  1. Golden Leaf Rd. 215, Mars Hill, 28754, USA

    Donald Armstrong

Rights and permissions

Reprints and permissions

Copyright information

© 2010 Humana Press, a part of Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Garrett, A.R., Murray, B.K., Robison, R.A., O’Neill, K.L. (2010). Measuring Antioxidant Capacity Using the ORAC and TOSC Assays. In: Armstrong, D. (eds) Advanced Protocols in Oxidative Stress II. Methods in Molecular Biology, vol 594. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-60761-411-1_17

Download citation

  • DOI: https://doi.org/10.1007/978-1-60761-411-1_17

  • Published:

  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-60761-410-4

  • Online ISBN: 978-1-60761-411-1

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation