Grzelakowska Aleksandra, Kalyanaraman Balaraman, Zielonka Jacek
Department of Biophysics, Medical College of Wisconsin, Milwaukee, United States.
Institute of Polymer and Dye Technology, Lodz University of Technology, Lodz, Poland.
Redox Biochem Chem. 2024 Dec;10. doi: 10.1016/j.rbc.2024.100034. Epub 2024 Jul 26.
Peroxynitrite (ONOO/ONOOH) is a short-lived but highly reactive species that is formed in the diffusion-controlled reaction between nitric oxide and the superoxide radical anion. It can oxidize certain biomolecules and has been considered as a key cellular oxidant formed under various pathophysiological conditions. It is crucial to selectively detect and quantify ONOO to determine its role in biological processes. In this review, we discuss various approaches used to detect ONOO in cell-free and cellular systems with the major emphasis on small-molecule chemical probes. We review the chemical principles and mechanisms responsible for the formation of the detectable products, and plausible limitations of the probes. We recommend the use of boronate-based chemical probes for ONOO, as they react directly and rapidly with ONOO, they produce minor but ONOO‒specific products, and the reaction kinetics and mechanism have been rigorously characterized. Specific experimental approaches and protocols for the detection of ONOO in cell-free, cellular, and systems using boronate-based molecular probes are provided (as shown in Boxes 1-6).
过氧亚硝酸盐（ONOO⁻/ONOOH）是一种寿命短暂但反应活性极高的物质，它在一氧化氮与超氧阴离子自由基的扩散控制反应中形成。它能够氧化某些生物分子，并且被认为是在各种病理生理条件下形成的关键细胞氧化剂。选择性地检测和定量过氧亚硝酸盐对于确定其在生物过程中的作用至关重要。在这篇综述中，我们讨论了用于在无细胞和细胞系统中检测过氧亚硝酸盐的各种方法，主要侧重于小分子化学探针。我们回顾了负责形成可检测产物的化学原理和机制，以及探针可能存在的局限性。我们推荐使用基于硼酸酯的化学探针来检测过氧亚硝酸盐，因为它们能直接且快速地与过氧亚硝酸盐反应，产生少量但针对过氧亚硝酸盐的特异性产物，并且反应动力学和机制已得到严格表征。本文还提供了使用基于硼酸酯的分子探针在无细胞、细胞及相关系统中检测过氧亚硝酸盐的具体实验方法和方案（如方框1 - 6所示）。
