一种新的产生和检测过氧亚硝基的化学发光体系

A novel chemiluminescence system for producing and measuring peroxynitrite anions

研究产生和消除过氧亚硝基(Peroxynitriteanion,ONOO?)是自由基生物学和医学的一项紧迫任务。为此,建立了一种新的产生和检测ONOO?的化学发光体系,并评估了检测的最佳条件。该方法为:10mmol/L羟胺与0.5mol/LNaOH、1mmol/L、乙二胺四乙酸二钠(Disodiumethylenediaminetetraacetate,EDTA)反应产生ONOO?,随后用MnO2粉末过滤反应液以除去H2O2,然后在测量管内加入100μL抗氧化剂和860μL0.1mol/L的碳酸盐缓冲液(pH10),再加入40μL过滤后的ONOO?反应液、混匀、延迟10s,测10s化学发光强度。以抗氧化剂茶多酚、抗坏血酸、芦丁和黄芩苷作清除ONOO?的实验。结果表明,本体系是一种灵敏价廉、可靠易行的筛选ONOO?清除剂的新方法。

The study for generating and scavenging peroxynitrite anions （ONOO^-） is an urgent task in free radical biology and medicine. Therefore, a novel chemiluminescence system for producing and measuring ONOO^- has been established. The optimal conditions for detecting ONOO^- were evaluated. The method can be described as follows. A mixture solution containing 10 mmo l / L hydroxylamine, 0.5 mo l / L NaOH and 1 mmo l / L EDTA was stirred vigorously in aerobic conditions for about 3 h to form ONOO^-. Certain amount of MnO2 powder was added to the mixture solution to eliminate H2O2 generated in the reaction. Then 100 μL antioxidant and 860μL 0.1mol/L carbonate buffer （pH10） were added into a test tube. By adding 40μL filtrated ONOO-solution into the tube, chemiluminescence （CL） reaction was initiated. Ten seconds after the solution was uniformly mixed, CL intensity of the reaction solution could be determined in a measurement of 10 seconds. The method was tested with tea polyphenol, ascorbic acid, rutin and baicalin, whish are known to have effects of scavenging ONOO^-. The tests show that the system is sensitive, low-cost, credible and easy method for screening ONOO^-.