The evaluation of new and isotopically labeled isoindoline nitroxides and an azaphenalene nitroxide for EPR oximetry

Abstract

Isoindoline nitroxides are potentially useful probes for viable biological systems, exhibiting low cytotoxicity, moderate rates of biological reduction and favorable Electron Paramagnetic Resonance (EPR) characteristics. We have evaluated the anionic (5-carboxy-1,1,3,3-tetramethylisoindolin-2-yloxyl; CTMIO), cationic (5-(N,N,N-trimethylammonio)-1,1,3,3-tetramethylisoindolin-2-yloxyl iodide, QATMIO) and neutral (1,1,3,3-tetramethylisoindolin-2-yloxyl; TMIO) nitroxides and their isotopically labeled analogues (2H12- and/or 2H12-15N-labeled) as potential EPR oximetry probes. An active ester analogue of CTMIO, designed to localize intracellularly, and the azaphenalene nitroxide 1,1,3,3-tetramethyl-2,3-dihydro-2-azaphenalen-2-yloxyl (TMAO) were also studied. While the EPR spectra of the unlabeled nitroxides exhibit high sensitivity to O2 concentration, deuteration resulted in a loss of superhyperfine features and a subsequent reduction in O2 sensitivity. Labeling the nitroxides with 15N increased the signal intensity and this may be useful in decreasing the detection limits for in vivo measurements. The active ester nitroxide showed approximately 6% intracellular localization and low cytotoxicity. The EPR spectra of TMAO nitroxide indicated an increased rigidity in the nitroxide ring, due to dibenzo-annulation.

In vivo EPR oximetry using an isotopically-substituted nitroxide: Potential for quantitative measurement of tissue oxygen

“…Therefore, nitroxide 1 is more O 2 -sensitive than nitroxide 3 and should be able to resolve smaller changes in O 2 . As expected, at 1 GHz, the spectral characteristics of nitroxides 1 and 3 , and their relative responsiveness to O 2 are comparable to our previous findings and for other similar spin labels at 9.5 GHz [26,27,30]. …”

First determination of the spin relaxation properties of a nitronyl nitroxide in solution by electron spin echoes at X-band: A comparison with Tempone

“…Among stable radicals, the two more popular classes are nitroxides and nitronyl nitroxides [11,12]; other stable radicals, as the triarylmethyl (trityl) ones, are used more rarely and are less suitable in some applications [13]. As the EPR spectrum of nitroxides is simpler with respect to that of nitronyl nitroxides, which is given by five major lines [14], only the former radicals are routinely used as spin probes.…”

Synthesis and Reduction Kinetics of Five Ibuprofen–Nitroxides for Ascorbic Acid and Methyl Radicals

“…Synthesis Six nitroxides (6)(7)(8)(9)(10)(11) were synthesized by the synthetic methods shown in Chart S1. Sterically shielded nitroxides 7, 10 and 11 were synthesized according to the method established by Yamada's group [4][5][6][7] and Rajca's group.…”

“…[7][8][9] Since ibuprofen-nitroxides were insoluble in PBS due to their lipophilicity, each nitroxide was added in a mixture of 1 mL of n-octanol and 1 mL of PBS (0.1 M, pH 7.4), and the resultant mixture was mixed vigorously for 1 h, and then the mixture was centrifuged at 3000 rpm for 5 min. Both n-octanol and PBS were subjected to the EPR measurement.…”

“…They had different reduction rates for reductants such as ascorbic acid (AsA), and these were applied as an invivo probe. [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17] The rate constant of reduction of a nitroxide probe in vitro is expected to be a guideline of its half-life in vivo. [9][10][11] The rate constants of them are greatly different and could not measure under the same conditions.…”