一种用于测量针对强效自由基反应的总抗氧化反应的新型自动化方法。

A novel automated method to measure total antioxidant response against potent free radical reactions.

作者信息

Erel Ozcan

机构信息

Clinical Biochemistry Department, Medical Faculty, Research Hospital, Harran University, Sanliurfa TR63200, Turkey.

出版信息

Clin Biochem. 2004 Feb;37(2):112-9. doi: 10.1016/j.clinbiochem.2003.10.014.

DOI:10.1016/j.clinbiochem.2003.10.014

PMID:14725941

Abstract

OBJECTIVES

Oxidative damage of biomolecules occurs as a result of potent free radical reactions. In this study, a novel, colorimetric and fully automated method for measuring total antioxidant response (TAR) against potent free radical reactions is described.

DESIGN AND METHODS

Potent free radical reactions were initiated with the production of hydroxyl radical (OH()) via Fenton reaction, and the rate of the reactions was monitored by following the absorbance of colored dianisidyl radicals. Ortho-dianisidine (10 mM) and ferrous ammonium sulfate (45 microM) were dissolved in KCl/HCl solution (75 mM, pH 1.8). This mixture was named as Reagent 1 and hydrogen peroxide solution (7.5 mM) as Reagent 2. The OH(), produced by mixing of R1 and R2, oxidized o-dianisidine molecules into dianisidyl radicals, leading to a bright yellow-brown color development within seconds. Antioxidants, present in the sample, suppressed the color formation to a degree that is proportional to their concentrations. The method was applied to an automated analyzer and analytical performance characteristics of the assay were determined.

RESULTS

Vitamin C and Trolox, reduced glutathione, bilirubin, uric acid and (+/-)-catechin solutions suppressed the color formation depending on their concentrations. Serum TAR against potent free radical reactions was lower in patients with chronic renal failure (1.13 +/- 0.21 mmol Trolox equiv./l) and was higher in the individuals with neonatal icterus (2.82 +/- 1.18 mmol Trolox equiv./l) than in healthy subjects (1.54 +/- 0.15 mmol Trolox equiv./l).

CONCLUSIONS

The easy, inexpensive and fully automated method described can be used to measure TAR of samples against potent free radical reactions.

摘要

目的

生物分子的氧化损伤是由强效自由基反应导致的。在本研究中，描述了一种用于测量针对强效自由基反应的总抗氧化反应（TAR）的新型比色全自动方法。

设计与方法

通过芬顿反应产生羟基自由基（OH()）引发强效自由基反应，并通过跟踪有色二茴香胺基自由基的吸光度来监测反应速率。将邻二茴香胺（10 mM）和硫酸亚铁铵（45 μM）溶解于KCl/HCl溶液（75 mM，pH 1.8）中。该混合物被命名为试剂1，过氧化氢溶液（7.5 mM）为试剂2。通过将R1和R2混合产生的OH()将邻二茴香胺分子氧化为二茴香胺基自由基，数秒内导致亮黄棕色显色。样品中存在的抗氧化剂会抑制颜色形成，其程度与它们的浓度成正比。该方法应用于自动分析仪并测定了该测定方法的分析性能特征。

结果

维生素C、Trolox、还原型谷胱甘肽、胆红素、尿酸和（±）-儿茶素溶液根据其浓度抑制颜色形成。慢性肾衰竭患者针对强效自由基反应的血清TAR较低（1.13±0.21 mmol Trolox当量/升），新生儿黄疸患者（2.82±1.18 mmol Trolox当量/升）的血清TAR高于健康受试者（1.54±0.15 mmol Trolox当量/升）。