Abstract
Non-structural phenolic compounds perform a variety of functions in plants, including acting as antioxidants. We describe a microplate-adapted colorimetric total phenolics assay that utilizes Folin–Ciocalteu (F–C) reagent. The F–C assay relies on the transfer of electrons in alkaline medium from phenolic compounds to phosphomolybdic/phosphotungstic acid complexes, which are determined spectroscopically at 765 nm. Although the electron transfer reaction is not specific for phenolic compounds, the extraction procedure eliminates approximately 85% of ascorbic acid and other potentially interfering compounds. This assay is performed in microcentrifuge tubes and assessed in a 96-well plate reader. At least 64 samples can be processed in 1 d.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 12 print issues and online access
$259.00 per year
only $21.58 per issue
Buy this article
- Purchase on SpringerLink
- Instant access to full article PDF
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
References
Croteau, R., Kutchan, T.M. & Lewis, N.G. Natural products (secondary metabolites). In Biochemistry & Molecular Biology of Plants (eds. Buchanan, B., Gruissem, W. & Jones, R.L.) 1250–1318 (American Society of Plant Physiologists, Beltsville, MD, USA, 2000).
Grace, S.C. & Logan, B.A. Energy dissipation and radical scavenging by the plant phenylpropanoid pathway. Phil. Trans. R. Soc. Lond. B 355, 1499–1510 (2000).
Grace, S.C. Phenolics as antioxidants. In Antioxidants and Reactive Oxygen Species in Plants (ed. Smirnoff, N.) 141–168 (Blackwell Publishing, Oxford, UK, 2005).
Bors, W., Heller, W., Michel, C. & Saran, M. Flavonoids as antioxidants: determination of radical-scavenging efficiencies. Methods Enzymol. 186, 343–355 (1990).
Bors, W., Michel, C. & Saran, M. Flavonoid antioxidants: rate constants for reactions with oxygen radicals. Methods Enzymol. 234, 420–429 (1994).
Dixon, R.A. & Paiva, N.L. Stress-induced phenylpropanoid metabolism. Plant Cell 7, 1085–1097 (1995).
Pasqualini, S. et al. Ozone-induced cell death in tobacco cultivar Bel W3 plants. The role of programmed cell death in lesion formation. Plant Physiol. 133, 1122–1134 (2003).
Robards, K. & Antolovich, M. Analytical chemistry of fruit bioflavonoids a review. Analyst 122, 11R–34R (1997).
Magalhães, L.M., Segundo, M.A., Reis, S., Lima, J.L. & Rangel, A.O. Automatic method for the determination of Folin–Ciocalteu reducing capacity in food products. J. Agric. Food Chem. 54, 5241–5246 (2006).
Folin, O. & Ciocalteu, V. On tyrosine and tryptophane determinations in proteins. J. Biol. Chem. 73, 627–650 (1927).
Singleton, V.L. & Rossi, J.A. Colorimetry of total phenolics with phosphomolybdic-phosphotungstic acid reagents. Am. J. Enol. Vitic. 16, 144–158 (1965).
Prior, R.L., Wu, X. & Schaich, K. Standardized methods for the determination of antioxidant capacity and phenolics in foods and dietary supplements. J. Agric. Food Chem. 53, 4290–4302 (2005).
Singleton, V.L., Orthofer, R. & Lamuela-Raventós,, R.M. Analysis of total phenols and other oxidation substrates and antioxidants by means of Folin-Ciocalteu reagent. Methods Enzymol. 299, 152–178 (1999).
Huang, D., Ou, B. & Prior, R.L. The chemistry behind antioxidant capacity assays. J. Agric. Food Chem. 53, 1841–1856 (2005).
Booker, F.L. & Miller, J.E. Phenylpropanoid metabolism and phenolic composition of soybean [Glycine max (L.) Merr.] leaves following exposure to ozone. J. Exp. Bot. 49, 1191–1202 (1998).
Gillespie, K.M., Chae, J.M. & Ainsworth, E.A. Rapid measurement of total antioxidant capacity in plants. Nat. Protoc. 2, 867–870 (2007).
Gillespie, K.M. & Ainsworth, E.A. Measurement of reduced, oxidized and total ascorbate content in plants. Nat. Protoc. 2, 871–874 (2007).
Acknowledgements
This research was supported by the Office of Science (BER), U.S. Department of Energy, Grant no. DE-FG02-04ER63849.
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Competing interests
The authors declare no competing financial interests.
Rights and permissions
About this article
Cite this article
Ainsworth, E., Gillespie, K. Estimation of total phenolic content and other oxidation substrates in plant tissues using Folin–Ciocalteu reagent. Nat Protoc 2, 875–877 (2007). https://doi.org/10.1038/nprot.2007.102
Published:
Issue Date:
DOI: https://doi.org/10.1038/nprot.2007.102
This article is cited by
-
Integrated metabolomics and transcriptomics analysis highlight key pathways involved in the somatic embryogenesis of Darjeeling tea
BMC Genomics (2024)
-
Multivariate investigation of Moringa oleifera morpho-physiological and biochemical traits under various water regimes
BMC Plant Biology (2024)
-
Variability in proximate composition, phytochemical traits and antioxidant properties of Iranian agro-ecotypic populations of fenugreek (Trigonella foenum-graecum L.)
Scientific Reports (2024)
-
Foliar application of pyroligneous acid acts synergistically with fertilizer to improve the productivity and phytochemical properties of greenhouse-grown tomato
Scientific Reports (2024)
-
Chitosan nanoparticles encapsulating curcumin counteract salt-mediated ionic toxicity in wheat seedlings: an ecofriendly and sustainable approach
Environmental Science and Pollution Research (2024)
Comments
By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.