Jacewicz Dagmara, Siedlecka-Kroplewska Kamila, Pranczk Joanna, Wyrzykowski Dariusz, Woźniak Michał, Chmurzyński Lech
Faculty of Chemistry, University of Gdańsk, Wita Stwosza 63, Gdańsk 80-308, Poland.
Department of Histology, Medical University of Gdańsk, Dębinki 1, Gdańsk 80-211, Poland.
Molecules. 2014 Jun 23;19(6):8533-43. doi: 10.3390/molecules19068533.
The purpose of this study was to examine the application of the coordinated cis-[Cr(C2O4)(pm)(OH)2]+ cation where pm denotes pyridoxamine, as a specific sensing ion for the detection of hydrogen peroxide (H2O2). The proposed method for H2O2 detection includes two key steps. The first step is based on the nonenzymatic decarboxylation of pyruvate upon reaction with H2O2, while the second step is based on the interaction of cis-[Cr(C2O4)(pm)(OH2)2]+ with the CO2 released in the previous step. Using this method H2O2 generated during glutamate-induced oxidative stress was detected in HT22 hippocampal cells. The coordination ion cis-[Cr(C2O4)(pm)(OH2)2]+ and the spectrophotometric stopped-flow technique were applied to determine the CO2 concentration in cell lysates, supernatants and cell-free culture medium. Prior to CO2 assessment pyruvate was added to all samples studied. Pyruvate reacts with H2O2 with 1:1 stoichiometry, and consequently the amount of CO2 released in this reaction is equivalent to the amount of H2O2.
本研究的目的是考察顺式-[Cr(C₂O₄)(吡哆胺)(OH)₂]⁺阳离子(其中吡哆胺用pm表示)作为检测过氧化氢(H₂O₂)的特定传感离子的应用。所提出的H₂O₂检测方法包括两个关键步骤。第一步基于丙酮酸与H₂O₂反应时的非酶促脱羧作用,而第二步基于顺式-[Cr(C₂O₄)(pm)(OH₂)₂]⁺与上一步释放的CO₂的相互作用。使用该方法在HT22海马细胞中检测了谷氨酸诱导的氧化应激过程中产生的H₂O₂。应用配位离子顺式-[Cr(C₂O₄)(pm)(OH₂)₂]⁺和分光光度法停流技术测定细胞裂解物、上清液和无细胞培养基中的CO₂浓度。在评估CO₂之前,向所有研究样品中加入丙酮酸。丙酮酸与H₂O₂以1:1的化学计量比反应,因此该反应中释放的CO₂量与H₂O₂量相当。