QUANTITATION OF EPR SPECTRA BY SPECTRAL SIMULATION

电子顺磁共振（EPR）自旋捕获 广泛用于检测瞬时自由基的形成 生物和化学系统。 该技术使用自旋陷阱 与自由基反应形成持久的自旋加合物， 可以直接用EPR检测。 在许多情况下，激进的 识别可以通过特定的自旋加合物来完成 自由基具有独特的光谱特征。 自旋捕获一般具有 以定性方式使用，因为自旋加合物通常也 浓度低和/或太不稳定而无法准确 量化。 良好的信噪比对于 准确定量，因为 EPR 谱必须积分两次 以获得样品中自由基的浓度。 我们开发了一种方法，通过该方法可以将信号较差的光谱 噪声是可以量化的。 该方法使用光谱模拟 生成原始频谱的无噪声表示，其中 然后可以通过积分进行定量分析。 浓度 可以确定复杂混合物中单个自旋加合物的数量。 在 此外，还可以分析自由基产生的动力学，作为信号 不需要采用平均。 这种方法已被用于 确定黄嘌呤/黄嘌呤形成超氧化物的动力学 氧化酶和一氧化氮合酶。

Electron paramagnetic resonance (EPR) spin-trapping has been extensively used to detect the formation of transient free radicals in biological and chemical systems. The technique uses a spin-trap that reacts with the free radical to form a persistent spin-adduct, which can be detected directly by EPR. In many cases, radical identification can be accomplished as the spin-adduct of a particular radical has unique spectral features. Spin-trapping has generally been used in a qualitative manner as the spin-adducts are usually too low in concentration and/or too unstable to allow accurate quantification. A good signal to noise ratio is essential for accurate quantification as the EPR spectrum has to be integrated twice in order to derive the concentration of free radical in the sample. We have developed a methodology whereby spectra with poor signal to noise can be quantified. This method uses a spectral simulation to generate a noise free representation of the ori gina l spectrum, which can then be quantitatively analyzed by integration. The concentration of individual spin adducts in complex mixtures can be determined. In addition the kinetics of radical production can be analyzed, as signal averaging need not be employed. This approach has been used to determine the kinetics of superoxide formation from xanthine/xanthine oxidase and nitric oxide synthase.