Ching Simon Y L, Hall Jon, Croft Kevin, Beilby John, Rossi Enrico, Ghisalberti Emilio
PathWest Laboratory Medicine WA, Queen Elizabeth II Medical Centre, Nedlands, Western Australia 6009, Australia.
Anal Biochem. 2006 Jun 15;353(2):257-65. doi: 10.1016/j.ab.2006.02.027. Epub 2006 Mar 15.
Few methods for assessing total antioxidant capacity (TAC) include both the percentage of inhibition and the length of inhibition in the measurement. Available methods require above ambient constant temperature incubation, reaction preheating, and/or separate assays for testing hydrophilic and hydrophobic samples. We describe a high-throughput method, antioxidant inhibition of oxygen radicals (AIOR), that overcomes these difficulties. AIOR uses peroxyl radicals to trigger a decrease in fluorescence of the indicator molecule, uroporphyrin I, which is delayed by the presence of antioxidants. The area under the curve is measured by a fluorescence spectrophotometer in a 96-well microplate format, and TAC results are expressed as millimole/liter Trolox equivalents. AIOR is performed at ambient temperature and is applicable to samples in either aqueous or common organic solvents. The reaction between uroporphyrin I and the peroxyl radicals generated from 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH) was found to be of first-order kinetics with a mean rate constant (k) of 0.0254. Applications to measure antioxidant capacity are demonstrated on individual chemicals and biological samples. The method has good linearity, within- and between-assay precision, and recovery.
很少有评估总抗氧化能力（TAC）的方法在测量中同时包含抑制百分比和抑制时长。现有方法需要高于环境温度的恒温孵育、反应预热以及/或者对亲水性和疏水性样品进行单独检测。我们描述了一种高通量方法——抗氧化剂对氧自由基的抑制作用（AIOR），该方法克服了这些困难。AIOR利用过氧自由基引发指示剂分子尿卟啉I的荧光下降，而抗氧化剂的存在会延迟这种下降。通过荧光分光光度计在96孔微孔板中测量曲线下面积，TAC结果以毫摩尔/升Trolox当量表示。AIOR在环境温度下进行，适用于水性或常见有机溶剂中的样品。发现尿卟啉I与2,2'-偶氮二异丁脒二盐酸盐（AAPH）产生的过氧自由基之间的反应为一级动力学，平均速率常数（k）为0.0254。在单个化学品和生物样品上展示了该方法在测量抗氧化能力方面的应用。该方法具有良好的线性、批内和批间精密度以及回收率。
