Journal
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
Volume 65, Issue 30, Pages 6288-6297Publisher
AMER CHEMICAL SOC
DOI: 10.1021/acs.jafc.7b02247
Keywords
antioxidant assay; PTIO center dot scavenging; oxygen-centered radical; analytical method; 2-phenyl-4,4,5,5-tetramethylimidazoline-l-oxyl 3-oxide; H+ transfer
Funding
- National Natural Science Foundation of China [81573558]
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Current in vitro antioxidant assays have several limitations, which frequently cause inconsistent results. The study develops a new antioxidant assay using the 2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide radical (PTIO center dot). After the investigation of various factors, the experimental protocol was briefly recommended as follows: PTIO center dot and the sample solution were added to phosphate buffer (pH 7.4, 50 mM), incubated at 37 degrees C for 2 h, and then spectrophotometrically measured at 557 nm. The validation test based on 20 pure compounds and 30 lyophilized aqueous extracts suggested that PTIO center dot scavenging had a good linear relationship, stability, and reproducibility. In the ultra-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight tandem mass spectrometry analysis, PTIO center dot was observed to give m/z 234 when encountering L-ascorbic acid. As an antioxidant assay, PTIO center dot scavenging possesses four advantages, i.e., oxygen-centered radical, physiological aqueous solution, simple and direct measurement, and less interference from the tested sample. It can also satisfactorily analyze the antioxidant structure-activity relationship. PTIO center dot scavenging has no stereospecificity and is at least involved in H+ transfer.
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