Metal ions and their interactions with biomolecules play an important role in human health. However, optical detectors commonly used for HPCE cannot directly detect metal ions without UV absorption. To make up for the shortcomings of existing HPCE detectors, a new universal HPCE detection system called an interface-induced current detector (IICRD) was constructed previously, with no need for derivatization procedures or complex instrumental modifications. Meanwhile, most of the reported studies on metal–biomolecule interactions only focused on the detection and analysis of biomolecules, commonly causing inaccurate or false-negative results, which is yet to be resolved. Here, the application of HPCE-IICRD realized the determination of metal–biomolecule interactions by directly measuring the electrophoretic parameters of metal ions for the first time, indicating that the interaction intensity can be measured more directly and accurately. Furthermore, an improved affinity capillary electrophoresis (ACE) method called relative mobility shift partial filling ACE-IICRD (rmsPF-ACE-IICRD) was originally developed to quantitatively analyze the binding strength. Binding behaviors between twelve free metal ions and three types of biomolecules (including two blood proteins, two enzyme proteins and two native DNAs) were investigated, and the values of the equilibrium dissociation constant (K_D) of metal–biomolecule complexes were calculated and evaluated by the nonlinear chromatography (NLC) method. The experimental results were basically consistent with the literature values. In particular, heavy metal ions showed stronger interactions with proteins and enzymes, while metal ions tended to show stronger binding with native DNAs than proteins and enzymes, which were in agreement with literature results. The combined use of HPCE-IICRD and rmsPF-ACE showed great advantages such as no need for pretreatment, low operating cost, good repeatability, simple operation and no interference from coexisting substances, which is hopeful to become an efficient metal ion detection method and also to expand the application scope of IICRD in the future.

中文翻译：

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通过相对迁移率偏移部分填充亲和毛细管电泳确定金属-生物分子相互作用


金属离子及其与生物分子的相互作用对人类健康起着重要作用。然而，通常用于 HPCE 的光学检测器无法在没有紫外线吸收的情况下直接检测金属离子。为了弥补现有 HPCE 检测器的缺点，之前构建了一种新的通用 HPCE 检测系统，称为界面感应电流检测器 （IICRD），无需衍生化程序或复杂的仪器修改。同时，大多数已报道的关于金属-生物分子相互作用的研究仅集中在生物分子的检测和分析上，这通常会导致不准确或假阴性的结果，这尚未得到解决。在这里，HPCE-IICRD 的应用首次通过直接测量金属离子的电泳参数实现了金属-生物分子相互作用的测定，表明可以更直接、更准确地测量相互作用强度。此外，最初开发了一种改进的亲和毛细管电泳 （ACE） 方法，称为相对迁移率偏移部分填充 ACE-IICRD （rmsPF-ACE-IICRD） 来定量分析结合强度。研究了 12 种游离金属离子与 3 种生物分子 （包括 2 种血液蛋白、2 种酶蛋白和 2 种天然 DNA） 之间的结合行为，并通过非线性色谱 （NLC） 方法计算和评价金属-生物分子复合物的平衡解离常数 （K_D） 值。实验结果与文献值基本一致。 特别是，重金属离子与蛋白质和酶的相互作用更强，而金属离子与天然 DNA 的结合往往比蛋白质和酶更强，这与文献结果一致。HPCE-IICRD 和 rmsPF-ACE 的联合使用显示出无需预处理、运行成本低、重复性好、操作简单、不受共存物质干扰等巨大优势，有望成为一种高效的金属离子检测方法，也为未来扩大 IICRD 的应用范围。