Spectrophotometric Assay for Superoxide Dismutase Based on the Reduction of Highly Water-soluble Tetrazolium Salts by Xanthine-Xanthine Oxidase
Abstract
Two novel highly water-soluble tetrazolium salts, WST-1 (4-[3-(4-iodophenyl)-2-(4-nitrophenyl)-2H-5-tetrazolio]-1,3-benzene disulfonate sodium salt) and WST-8 (4-[3-(2-methoxy-4-nitrophenyl)-2-(4-nitrophenyl)-2H-5-tetrazolio]-1,3-benzene disulfonate sodium salt) were applied to the assay of superoxide dismutase (SOD). The superoxide anion generated by xanthine/xanthine oxidase (XO) reduced WST-1 and WST-8 to water-soluble formazans which exhibited absorbance maxima at 438 and 460 nm, respectively. The rates of reduction were linearly related to the XO activity, and reduction was inhibited by SOD. Complete inhibition by SOD of the reduction of both WST-1 and WST-8 was achieved, suggesting that these WSTs were not reduced with XO. WST-1 was found more useful than WST-8 because it had shown higher sensitivity which was apparently not dependent on the assay pH value in the range pH 8.0-10.2. These properties of WST-1 are ideal for the spectrophotometric assay of SOD in an aqueous system.
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Superoxide ions are generated from the conversion of xanthine and O2 to uric acid and H2O2 by xanthine oxidase.
“Superoxide ions are generated from the conversion of xanthine and O2 to uric acid and H2O2 by xanthine oxidase (XO).”
From Superoxide Anion Chemistry—Its Role at the Core of the Innate Immunity (2023) · cites this paper as [106]
Superoxide anion converts WST-1 to the coloured product WST-1 formazan.
“The superoxide anion then converts the tetrazolium salt WST-1 to the coloured product WST-1 formazan [106], Figure 41.”
From Superoxide Anion Chemistry—Its Role at the Core of the Innate Immunity (2023) · cites this paper as [106]
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Superoxide Anion Chemistry—Its Role at the Core of the Innate Immunity
Andrés, C. M. C., Pérez de la Lastra, J. M., Andrés Juan, C., Plou, F. J., & Pérez-Lebeña, E. (2023) · International Journal of Molecular Sciences · doi:10.3390/ijms24031841 · Cited by 342 · see citing works
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- Authors
- UKEDA, H., KAWANA, D., MAEDA, S., & SAWAMURA, M.
- Affiliations
- University of Kochi (JP)
- Venue
- Bioscience, Biotechnology, and Biochemistry
- Publisher
- Oxford University Press (OUP)
- Published
- 1999-01
- Volume
- 63
- Issue
- 3
- Pages
- 485-488
- ISSN
- 0916-8451 (ISSN-L)
- 1347-6947
- OpenAlex
- W2043512927
- Cited by
- 263
- References
- 12
- Citations by year
- 2026: 27 · 2025: 23 · 2024: 25 · 2023: 23 · 2022: 13 · 2021: 19 · 2020: 5 · 2019: 13 · 2018: 8 · 2017: 5 · 2016: 7 · 2015: 3 · 2014: 8 · 2013: 8 · 2012: 6
- FWCI
- 1.03
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- Bioscience Biotechnology and Biochemistry (publishedVersion)
- PubMed (publishedVersion)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Redox biology and oxidative stress
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Cancer and biochemical research
- Health Sciences › Medicine › Biochemistry › Antioxidant Activity and Oxidative Stress
- Keywords
- superoxide dismutase, tetrazolium salts, xanthine/xanthine oxidase, superoxide, formazan reduction, spectrophotometric assay, enzyme inhibition
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