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首页> 美国卫生研究院文献>International Journal of Molecular Sciences >Function of Hemoglobin-Based Oxygen Carriers: Determination of Methemoglobin Content by Spectral Extinction Measurements
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Function of Hemoglobin-Based Oxygen Carriers: Determination of Methemoglobin Content by Spectral Extinction Measurements

机译：基于血红蛋白的氧载体的功能：通过光谱消光测量测定甲基酚蛋白含量

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Suspensions of hemoglobin microparticles (HbMPs) are promising tools as oxygen therapeutics. For the approval of clinical studies extensive characterization of these HbMPs with a size of about 750 nm is required regarding physical properties, function, pharmaco-kinetics and toxicology. The standard absorbance measurements in blood gas analyzers require dissolution of red blood cells which does not work for HbMP. Therefore, we have developed a robust and rapid optical method for the quality and functionality control of HbMPs. It allows simultaneous determination of the portion of the two states of hemoglobin oxygenated hemoglobin (oxyHb) and deoxygenated hemoglobin (deoxyHb) as well as the content of methemoglobin (metHb). Based on the measurement of collimated transmission spectra between 300 nm and 800 nm, the average extinction cross section of HbMPs is derived. A numerical method is applied to determine the composition of the HbMPs based on their wavelength-dependent refractive index (RI), which is a superposition of the three different states of Hb. Thus, light-scattering properties, including extinction cross sections can be simulated for different compositions and sizes. By comparison to measured spectra, the relative concentrations of oxyHb, deoxyHb, metHb are accessible. For validation of the optically determined composition of the HbMPs, we used X-ray fluorescence spectrometry for the ratio of Fe(II) (oxyHb/deoxyHb) and Fe(III) (metHb). High accuracy density measurements served to access heme-free proteins, size was determined by dynamic light scattering and analytical centrifugation and the shape of the HbMPs was visualized by electron and atomic force microscopy.
机译：血红蛋白微粒（HBMPS）的悬浮液是有前途的工具作为氧气治疗方法。对于临床研究的批准，对于物理性质，功能，药物动力学和毒理学，需要大尺寸的临床研究的广泛表征这些HBMP的尺寸约为750nm。血气分析仪中的标准吸光度测量需要溶解红细胞，其不适用于HBMP。因此，我们开发了一种强大而快速的光学方法，用于HBMP的质量和功能控制。它允许同时测定血红蛋白含氧血红蛋白（氧气）和脱氧血红蛋白（DEOXYHB）的两种状态以及甲基酚（甲基溴）的含量。基于300nm和800nm之间的准直透射光谱的测量，导出HBMP的平均消光横截面。应用数值方法以基于其波长依赖性折射率（RI）来确定HBMP的组成，这是HB的三种不同状态的叠加。因此，可以模拟不同的组合物和尺寸的光散射性能，包括消光横截面。通过比较测量光谱，可获得氧气，脱氧，甲基甲基的相对浓度。为了验证HBMPS的光学确定的组合物，我们使用X射线荧光光谱法用于Fe（II）（氧气/脱氧，Fe（III）（甲基）的比例。高精度密度测量用于接入无血红素蛋白质，尺寸通过动态光散射测定，并通过电子和原子力显微镜观察HBMP的形状。

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