Nitro-Triarylmethyl Radical as Dual Oxygen and Superoxide Probe

Superoxide radical is involved in numerous physiological and pathophysiological processes. Tetrathiatriarylmethyl (TAM) radicals are knows to react with superoxide allowing measurement of superoxide production in biological media. We report the synthesis of a Nitro conjugated TAM radical showing a rate constant of 7 × 105 M−1s−1 which is two order of magnitude higher than other TAMs allowing high sensitivity measurement of superoxid

Dioxygen Binding and Sensing Proteins

Oxygen binding proteins (O2BIP) have been actively investigated over the last five decades due to their rich redox chemistry and function as O2 carriers in blood cells, as well as their function as gasotransmitters and sensors that modulate cellular signaling. A series of meetings on the periodic advances in the knowledge gained in the field of globin structure and function are conducted typically on a biannual basis. In the fall of 2018, the XXth International Conference was conducted, and very important papers with breakthrough discoveries were presented and very enthusiastically discussed. This was yet another highly successful meeting in the series. Select papers from this meeting were recently reviewed, updated and published over several issues of Antioxidants and Redox Signaling (ARS), as forum papers communicating the latest advances in this important area of redox biology. This forum editorial introduces these articles, and highlights their scientific significance in advancing the field. Each of these articles grew out of lectures presented in the meeting, and appears either as an original contribution or a comprehensive review in the journal. Overall, the articles published in the forum provide in-depth details on the recent developments in the field as well as point the way to future directions. These forum papers thus serve as an important summary of progress and the ongoing direction of this field, and serve to highlight recent advances in our understanding of O2BIP

Electron paramagnetic resonance evidence that cellular oxygen toxicity is caused by the generation of superoxide and hydroxyl free radicals

AbstractCells require molecular oxygen for the generation of energy through mitochondrial oxidative phosphorylation; however, high concentrations of oxygen are toxic and can cause cell death. A number of different mechanisms have been proposed to cause cellular oxygen toxicity. One hypothesis is that reactive oxygen free radicals may be generated; however free radical generation in hyperoxic cells has never been directly measured and the mechanism of this radical generation is unknown. In order to determine if cellular oxygen toxicity is free radical mediated, we applied electron paramagnetic resonance, EPR, spectroscopy using the spin trap 5,5′-dimethyl-1-pyrroline-N-oxide, DMPO, to measure free radical generation in hyperoxic pulmonary endothelial cells. Cells in air did not give rise to any detectable signal. However, cells exposed to 100% O2 for 30 min exhibited a prominent signal of trapped hydroxyl radical, DMPO-OH, while cell free buffer did not give rise to any detectable radical generation. This cellular radical generation was demonstrated to be derived from the superoxide radical since the observed signal was totally quenched by superoxide dismutase, but not by equal concentrations of the denatured enzyme. It was confirmed that the hydroxyl radical was generated since in the presence of ethanol the CH3·CH(OH) radical was formed. Loss of cell viability as measured by uptake of trypan blue dye was observed paralleling the measured free radical generation. Thus, superoxide and hydroxyl radicals are generated in hyperoxic pulmonary endothelial cells and this appears to be an important mechanism of cellular oxygen toxicity

Oxygen Regulates The Effective Diffusion Distance of Nitric Oxide in The Aortic Wall

Endothelium-derived nitric oxide (NO) is critical in maintaining vascular tone. Accumulating evidence shows that NO bioavailability is regulated by oxygen concentration. However, it is unclear to what extent the oxygen concentration regulates NO bioavailability in the vascular wall. In this study, a recently developed experimental setup was used to measure the NO diffusion flux across the aortic wall at various oxygen concentrations. It was observed that for a constant NO concentration at the endothelial surface, the measured NO diffusion flux out of the adventitial surface at [O2] = 0 μM is around fivefold greater than at [O2] = 150 μM, indicating that NO is consumed in the aortic wall in an oxygen-dependent manner. Analysis of experimental data shows that the rate of NO consumption in the aortic wall is first order with respect to [NO] and first order with respect to [O2], and the rate constant k1 was determined as (4.0 ± 0.3) × 103 M−1 s−1. Computer simulations demonstrate that NO concentration distribution significantly changes with oxygen concentration and the effective NO diffusion distance at low oxygen level ([O2] ≤ 25 μM) is significantly longer than that at high oxygen level ([O2] = 200 μM). These results suggest that oxygen-dependent NO consumption may play an important role in dilating blood vessels during hypoxia by increasing the effective NO diffusion distance

Structural factors controlling the spin-spin exchange coupling: EPR spectroscopic studies of highly asymmetric trityl-nitroxide biradicals

Highly asymmetric exchange-coupled biradicals, like the trityl-nitroxides (TN), possess particular magnetic properties opening new possibilities for their application in biophysical, physicochemical and biological studies. In the present work, we investigated the effect of the linker length on the spin-spin coupling interaction in TN biradicals using the newly synthesized biradicals CT02-GT, CT02-AT, CT02-VT and CT02-PPT as well as the previously reported biradicals TNN14 and TN1. Results show that the magnitude of the spin-spin interaction (J) can be easily tuned from ~ 4 G (conformer 1 in CT02-PPT) to over 1200 G (in TNN14) using various linkers separating the two radical moieties and with varying temperature. Computer simulation of EPR spectra was carried out to directly estimate J values of the TN biradicals. In addition to the spin-spin coupling interaction of TN biradicals, their g, hyperfine splitting and zero-field splitting interactions were explored at low temperature (220 K). Our present study clearly shows that the spin-spin interaction variation as a function of linker distance and temperature provides an effective strategy to develop new TN biradicals which can find wide applications in relevant fields