量研学術機関リポジトリ「QST-Repository」は、国立研究開発法人 量子科学技術研究開発機構に所属する職員等が生み出した学術成果（学会誌発表論文、学会発表、研究開発報告書、特許等）を集積しインターネット上で広く公開するサービスです。 Welcome to QST-Repository where we accumulates and discloses the academic research results(Journal Publications, Conference presentation, Research and Development Report, Patent, etc.) of the members of National Institutes for Quantum Science and Technology.
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Background: The radical-scavenging reaction of antioxidants is known to be significantly affected by pH, solvents, and metal ions. 2,2-Diphenyl-1-picrylhydrazyl (DPPH) radical has been used to evaluate the radical-scavenging activity of antioxidants as a reactivity model of reactive oxygen species. However, the insolubility of the DPPH radical in water has precluded its use in aqueous media. Recently, we have succeeded in solubilizing the DPPH radical in water using β-cyclodextrin (β-CD). This enables us to evaluate the scavenging activity of the DPPH radical by antioxidants in aqueous media. We report herein the effect of reaction media on the scavenging reaction of the DPPH radical by ascorbic acid, a representative water-soluble antioxidant.
Methods: 25 mL of boiling water (Milli-Q) was added to the mixture containing the DPPH radical (0.15 mmol) and β-CD (0.70 mmol), and the suspension was cooled to room temperature. The filtration of the suspension by a membrane filter (pore size: 0.22 µm) gave a deep violet solution of the DPPH radical. The rates of the scavenging reaction of the DPPH radical were determined by monitoring the absorbance change due to the DPPH radical using a stopped-flow technique on a UNISOKU RSP-1000-02NM spectrophotometer.
Results: The second-order rate constant (k) of the reaction between ascorbic acid and the β-CD-solubilized DPPH radical was determined to be 5.4 × 103 M–1 s–1 in phosphate buffer (0.05 M, pH 7.0) at 25 °C. The k value increased with increasing the pH value. Furthermore, magnesium perchlorate significantly accelerated this reaction. On the other hand, when water was replaced by deuterium oxide to prepare the phosphate buffer (0.05 M, pD 7.0), the k value significantly decreased to be 1.5 × 103 M–1 s–1.
Conclusion: The scavenging reaction of the DPPH radical by ascorbic acid is significantly affected by reaction media.
Effect of Reaction Media on the Scavenging Reaction of Water-Solubilized 2,2-Diphenyl-1-picrylhydrzyl Radical by Ascorbic Acid
