Abstract：
Glutathione (GSH) is indispensable for maintaining redox homeostasis in biological fluids and serves as a key component in cellular defense mechanisms. Accurate assessment of GSH relative to its oxidized counterpart, glutathione disulfide (GSSG), is critical for the early diagnosis and understanding of conditions related to oxidative stress. Despite existing methods for their quantification, the label-free and simultaneous measurement of GSH and GSSG in biological fluid presents significant challenges. Herein, we report the use of an alpha-hederin (Ah) nanopore for the direct measurement of the GSH:GSSG ratio in simulated biological fluid, containing fetal bovine serum (FBS). This system hinges on detecting characteristic relative ion blockades (ΔI/Io) as GSH and GSSG molecules pass through the Ah nanopore under an applied electric field. The distinct current blockage signals derived from the translocation of GSH and GSSG enabled us to determine the molar ratio of GSH and its oxidized form. Notably, the interactions between the hydroxyl groups of the sugar moiety lining the nanopore\'s inner surface and the sulfhydryl group of GSH significantly influence the translocation dynamics, resulting in a longer translocation time for GSH compared to GSSG. The Ah nanopore technology proposed in this study offers a promising approach for real-time, single molecule-level monitoring of glutathione redox status in biological fluids, eliminating the need for labeling or extensive sample preparation.
摘要：
谷胱甘肽（GSH）对于维持生物体液中的氧化还原稳态是必不可少的，并且是细胞防御机制的关键组成部分。GSH相对于其氧化对应物的准确评估，谷胱甘肽二硫化物（GSSG），对于早期诊断和了解与氧化应激相关的疾病至关重要。尽管现有的量化方法，生物流体中GSH和GSSG的无标记和同时测量提出了重大挑战。在这里，我们报告了使用α-hederin（Ah）纳米孔直接测量模拟生物流体中的GSH：GSSG比率，含胎牛血清（FBS）。当GSH和GSSG分子在施加的电场下通过Ah纳米孔时，该系统取决于检测特征相对离子阻塞（ΔI/Io）。来自GSH和GSSG易位的不同电流阻断信号使我们能够确定GSH及其氧化形式的摩尔比。值得注意的是,纳米孔的内表面的糖部分的羟基与GSH的巯基之间的相互作用显着影响易位动力学，与GSSG相比，GSH的易位时间更长。本研究中提出的Ah纳米孔技术为实时，生物液中谷胱甘肽氧化还原状态的单分子水平监测，消除了标签或大量样品制备的需要。
