Measurement of reduced, oxidized and total ascorbate content in plants

Kelly M. Gillespie, Elizabeth A. Ainsworth

Research output: Contribution to journalArticlepeer-review


Ascorbate is one of the major antioxidant metabolites in plant tissues. This protocol describes a microplate-adapted colorimetric ascorbate assay, in which ferric ion is reduced by ascorbate to the ferrous ion. The ferrous ion reacts with α-ά-bipyridl to form a complex with characteristic absorbance at 525 nm. With the chemical reduction of any dehydroascorbate (DHA) in a sample, total ascorbate can be assayed using the α-ά-bipyridl method, and DHA can be estimated by subtracting the reduced portion from the total ascorbate pool. The assay is performed in microcentrifuge tubes and assessed in a 96-well plate reader. Reduced ascorbate, DHA and total ascorbate of at least 64 experimental samples can be analyzed easily in 1 d.

Original languageEnglish (US)
Pages (from-to)871-874
Number of pages4
JournalNature Protocols
Issue number4
StatePublished - Apr 2007

ASJC Scopus subject areas

  • General Biochemistry, Genetics and Molecular Biology


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