芳环羟基化荧光法检测类Fenton反应产生的·OH

Fluorimetry Assay of Hydroxyl Radical by Fenton-Like Reaction-Aromatic Hydroxylation System

采用L-苯丙氨酸为探针,使用液相色谱仪(HPLC)荧光检测器检测了7种过渡金属离子参与类Fen-ton反应产生.OH的能力.试验采用的激发波长277nm,发射波长306nm.产物L-Tyr在反应前后的荧光变化可间接反映.OH产生量.实验结果表明Cr(Ⅲ)、Fe(Ⅱ)、Co(Ⅱ)、Cu(Ⅱ)、Pb(Ⅱ)产生.OH的能力较强,而Ni(Ⅱ)和Cd(Ⅱ)产生.OH的能力极弱..OH清除剂脱铁敏或甘露醇,能明显抑制产物L-Tyr的荧光强度变化,说明.OH的产生受到抑制,结果表明HPLC方法准确可靠.使用分光光度法对上述离子参与类Fenton反应产生.OH的实验进行了验证,得到了基本一致的结果,但芳环羟基化荧光法的结果更稳定,灵敏度更高,故此方法对于.OH检测以及抗.OH机理和应用研究等方面具有较高应用价值.

In this study, RP-HPLC with fluorescence detection（excitation 277 nm and emission 306 nm）was used to detect hydroxyl radical generated by Fenton like reaction system participatated by seven transition metal-ions, using a fluorescence probe, L-phenylalanine, which could be converted to highly fluorescent L-tyrosine in the generation of oOH the variety of fluorescence intensity reflected the generation of · OH. The experimental results indicated that ability of Cr（ Ⅲ ）,re（ Ⅱ ）,Co（ Ⅱ ）,Cu（ Ⅱ ）,Pb（ Ⅱ ） to produce ·OH is relatively, strong,but the ability of Cd（Ⅱ） and Ni（Ⅱ） is poor. The intensity of fluorescence in the systems could be inhibited obviously by mannite and desferrioxamine（DFO）,which proved ·OH was cleard. We used the UV spectrometer and got the same results. Aromatic hydroxylation was more effective and reliable. So this sensitive method can be applied to study the hydroxyl radical and hydroxyl radical scavenger.