Che P, Xu J, Shi H, Ma Y
Division of Science, Northeast Missouri State University, Kirksville 63501, USA.
J Chromatogr B Biomed Appl. 1995 Jul 7;669(1):45-51. doi: 10.1016/0378-4347(95)00041-g.
A capillary electrophoretic (HPCE) method that can be used to quantitatively determine trace amounts of iron has been developed and applied to determine the iron level in human serum. After precipitation of serum proteins, Fe(III) in the serum is reduced to Fe(II) with hydroxylamine hydrochloride, and a stable Fe(II)-1,10-phenanthroline complex is formed by adding 1,10-phenanthroline to the supernatant containing 2.5 mM ammonium acetate-acetic acid at pH 5.0. The Fe(II)-1,10-phenanthroline complex, [Fe(C12H8N2)3]2+, has a very strong absorbance at 270 nm (with a molar absorptivity of approximately 9.2.10(4)). By measuring the absorbance of [Fe(C12H8N2)3]2+ at 270 nm, the iron level in human serum can be precisely quantified. The interference from copper, a major interference in serum, can be totally eliminated due to the complete separation of [Fe(C12H8N2)3]2+ and the Cu(II)-1,10-phenanthroline complex. In addition, other problems that usually occurred with conventional spectrophotometric methods, such as co-precipitation and occlusion of iron during sample pretreatment, are significantly minimized due to the ability to wash the precipitate and the higher detection sensitivity. With this method, a single drop (10 microliters) of serum would be sufficient to determine the serum iron concentration. The method is reliable, sensitive, rapid and reproducible. Thus it is highly suitable for use in the clinical laboratory.
已开发出一种可用于定量测定痕量铁的毛细管电泳（HPCE）方法，并将其应用于测定人体血清中的铁含量。血清蛋白沉淀后，血清中的Fe(III)用盐酸羟胺还原为Fe(II)，在pH 5.0含有2.5 mM乙酸铵 - 乙酸的上清液中加入1,10 - 菲啰啉，形成稳定的Fe(II)-1,10 - 菲啰啉络合物。Fe(II)-1,10 - 菲啰啉络合物[Fe(C₁₂H₈N₂)₃]²⁺在270 nm处有很强的吸光度（摩尔吸光系数约为9.2×10⁴）。通过测量[Fe(C₁₂H₈N₂)₃]²⁺在270 nm处的吸光度，可精确量化人体血清中的铁含量。由于[Fe(C₁₂H₈N₂)₃]²⁺与Cu(II)-1,10 - 菲啰啉络合物能完全分离，血清中主要干扰物铜的干扰可被完全消除。此外，由于能够洗涤沉淀以及检测灵敏度更高，传统分光光度法通常出现的其他问题，如样品预处理过程中铁的共沉淀和包藏现象，也能显著减少。用这种方法，一滴（10微升）血清就足以测定血清铁浓度。该方法可靠、灵敏、快速且可重复。因此，它非常适合用于临床实验室。
