Spin Trapping Agent Research Articles

Isomeric sp2-C-conjugated porous organic polymer-mediated photo- and sono-catalytic detoxification of sulfur mustard simulant under ambient conditions

Isomeric sp2-C-conjugated porous organic polymer-mediated photo- and sono-catalytic detoxification of sulfur mustard simulant under ambient conditions

Pro-oxidant activity of biocompatible catechin stabilizer during photooxidation of polyolefins

Polymer plaques made of high-density polyethylene (HDPE) or cycloolefin copolymer (COC) were stabilized with natural phenolic stabilizer (+)-catechin (CAT) and aged using accelerated weathering technique (WOM; accelerated photooxidation). The efficiency of the phenolic stabilizer was compared with a well-established hindered amine stabilizer (HAS) Tinuvin®770 (Tin770). In pursuit of the stabilization and identification of short-living radicals generated in the process of photooxidation of the polymers, a spin trapping agent 2,4,6-tri-tert-butylnitrosobenzene (TTBNB) was added to selected samples. Profiles of oxidation products and crystallinity inside the HDPE plaques were determined by IR microspectroscopy, oxidation products in COC copolymers were identified using ATR spectroscopy, concentration profiles of radicals generated inside polymer plaques during WOM exposure were determined by ESRI, the changes of local mechanical properties at the surface of polymer plaques were characterized using microindentation hardness testing (MHI), and the surface morphology changes were studied by light and/or scanning electron microscopy. All methods are in accordance with the conclusion that the natural phenolic stabilizer CAT exhibited pro-oxidant activity during accelerated photooxidation of both polymers, i.e. higher concentration of oxidation products, bigger changes of local mechanical properties and/or more microcracks on the exposed surfaces in comparison with non-stabilized systems and systems stabilized with Tin770.

Biochemical and Mutational Analysis of Radical S-Adenosyl-L-Methionine Adenosylhopane Synthase HpnH from Zymomonas mobilis Reveals that the Conserved Residue Cysteine-106 Reduces a Radical Intermediate and Determines the Stereochemistry.

Adenosylhopane is a crucial precursor of C35 hopanoids, which are believed to modulate the fluidity and permeability of bacterial cell membranes. Adenosylhopane is formed by a crosslinking reaction between diploptene and a 5'-deoxyadenosyl radical that is generated by the radical S-adenosyl-L-methionine (SAM) enzyme HpnH. We previously showed that HpnH from Streptomyces coelicolor A3(2) (ScHpnH) converts diploptene to (22R)-adenosylhopane. However, the mechanism of the stereoselective C-C bond formation was unclear. Thus, here, we performed biochemical and mutational analysis of another HpnH, from the ethanol-producing bacterium Zymomonas mobilis (ZmHpnH). Similar to ScHpnH, wild-type ZmHpnH afforded (22R)-adenosylhopane. Conserved cysteine and tyrosine residues were suggested as possible hydrogen sources to quench the putative radical reaction intermediate. A Cys106Ala mutant of ZmHpnH had one-fortieth the activity of the wild-type enzyme and yielded both (22R)- and (22S)-adenosylhopane along with some related byproducts. Radical trapping experiments with a spin-trapping agent supported the generation of a radical intermediate in the ZmHpnH-catalyzed reaction. We propose that the thiol of Cys106 stereoselectively reduces the radical intermediate generated at the C22 position by the addition of the 5'-deoxadenosyl radical to diploptene, to complete the reaction.

A Critical Review on Removal of Gaseous Pollutants Using Sulfate Radical-based Advanced Oxidation Technologies.

Excessive emissions of gaseous pollutants such as SO2, NOx, heavy metals (Hg, As, etc.), H2S, VOCs, etc. have triggered a series of environmental pollution incidents. Sulfate radical (SO4•-)-based advanced oxidation technologies (AOTs) are one of the most promising gaseous pollutants removal technologies because they can not only produce active free radicals with strong oxidation ability to simultaneously degrade most of gaseous pollutants, but also their reaction processes are environmentally friendly. However, so far, the special review focusing on gaseous pollutants removal using SO4•--based AOTs is not reported. This review reports the latest advances in removal of gaseous pollutants (e.g., SO2, NOx, Hg, As, H2S, and VOCs) using SO4•--based AOTs. The performance, mechanism, active species identification and advantages/disadvantages of these removal technologies using SO4•--based AOTs are reviewed. The existing challenges and further research suggestions are also commented. Results show that SO4•--based AOTs possess good development potential in gaseous pollutant control field due to simple reagent transportation and storage, low product post-treatment requirements and strong degradation ability of refractory pollutants. Each SO4•--based AOT possesses its own advantages and disadvantages in terms of removal performance, cost, reliability, and product post-treatment. Low free radical yield, poor removal capacity, unclear removal mechanism/contribution of active species, unreliable technology and high cost are still the main problems in this field. The combined use of multiactivation technologies is one of the promising strategies to overcome these defects since it may make up for the shortcomings of independent technology. In order to improve free radical yield and pollutant removal capacity, enhancement of mass transfer and optimization design of reactor are critical issues. Comprehensive consideration of catalytic materials, removal chemistry, mass transfer and reactor is the promising route to solve these problems. In order to clarify removal mechanism, it is essential to select suitable free radical sacrificial agents, probes and spin trapping agents, which possess high selectivity for target specie, high solubility in water, and little effect on activity of catalyst itself and mass transfer/diffusion parameters. In order to further reduce investment and operating costs, it is necessary to carry out the related studies on simultaneous removal of more gaseous pollutants.

Synthesis and evaluation of cyclic nitrone derivatives as potential anti-cancer agents

Nitrones have been found to exhibit attractive biological values as immuno spin trapping agents. However, successful clinical cases of nitrone therapeutics are still lacking. Herein we report the synthesis and antiproliferative activity of a series of structurally diverse nitrone derivatives against a panel of 5 cancer cell lines, based on which indole- and pyrrole-fused were further evaluated by analogue preparation and in-vitro screening. Analogues with moderate to good potency were identified. This study shows the promise for further pursuit of nitrone-type small molecules in chemotherapy.

Imaging of Hydroxyl-Radical Generation Using Dynamic Nuclear Polarization-Magnetic Resonance Imaging and a Spin-Trapping Agent.

Reactive oxygen species (ROS) play an important role in cell metabolism, but they can cause oxidative damage to biomolecules. Among ROS, the hydroxyl radical (·OH) is one of the most reactive molecules in biological systems because of its high reaction rate constant. Therefore, imaging of ·OH could be useful for evaluation of the redox mechanism and diagnosis of oxidative diseases. In vivo dynamic nuclear polarization-magnetic resonance imaging (DNP-MRI) is a noninvasive imaging method to obtain spatiotemporal information about free radicals with MRI anatomical resolution. In this study, we investigated the visualization of hydroxyl radicals generated from the Fenton reaction by combining DNP-MRI with a spin-trapping agent (DMPO: 5,5-dimethyl-1-pyrroline N-oxide) for ·OH. Additionally, we demonstrated the radical-scavenging effect using four thiol-related reagents by DNP-MRI. We demonstrated that DNP enhancement could be induced by the DMPO-OH radical using the DNP-MRI/spin-trapping method and visualized ·OH generation for the first time. Maximum DNP enhancement was observed at an electron paramagnetic resonance irradiation frequency of 474.5 MHz. Furthermore, the radical-scavenging effect was simultaneously evaluated by the decrease in the DNP image value of DMPO-OH. An advantage of our methods is that they simultaneously investigate compound activity and the radical-scavenging effect.

Detection and identification of ethanal-derived spin-trapped free radicals using headspace thermal desorption gas chromatography-mass spectrometry (TD-GC-MS)

In this study, we demonstrate a novel approach to the detection and identification of the products of spin-trapped free radicals. Hydroxyl free radicals were generated by Fenton-based chemistry in the presence of ethanal and the spin-trapping agent N-tert-butyl-α-phenylnitrone (PBN). The resulting volatile compounds present in the reaction vial headspace were collected using thermal desorption (TD) and analysed by gas chromatography-mass spectrometry (GC-MS). Eleven compounds were detected in the headspace, and their identification was aided by using either a fluorinated or deuterated analogue of PBN as an alternative spin trap and/or deuterated ethanal (CD3CHO) as the secondary source of free radicals. The electron-ionisation (EI) mass spectra clearly demonstrate the “capture” of methyl radicals; two of the compounds detected were identified as containing one methyl group derived from ethanal, and four were shown to contain two methyl groups. This study demonstrates that sampling the reaction headspace using TD-GC-MS is a viable method for analysing products of free radical trapping, and potentially may be applied to a wide range of free radical systems.

Synthesis and Photocatalytic Properties of Titanium-Porphyrinic Aerogels.

We present novel titanium-porphyrinic gels (TPGs) and titanium-porphyrinic aerogels (TPAs), in which porphyrinic ligand tetrakis(4-carboxyphenyl)porphyrin is coordinated to Ti-oxo clusters. These hierarchically porous TPAs, with micro-, meso-, and macropores and reactant-concentration-dependent Brunauer-Emmett-Teller surface areas of 407-738 m2 g-1 , are prepared by CO2 critical point drying of TPGs. Although the Ti4+ → Ti3+ photoreduction of TPAs is less efficient than that of crystalline microporous Ti-porphyrinic framework DGIST-1, prompt diffusion of O2 and spin-trapping agents into the TPA pores causes the rapid generation of reactive oxygen species (ROS), as observed by EPR spectroscopy. When used as an ROS scavenger, large 1,3-diphenylisobenzofuran is degraded by the best-performing TPA 10 times faster than by DGIST-1, suggesting that the accessibility of molecules (reactants) to pores (reactive centers) strongly influences photocatalytic activity.

Pentacyanoferrate(II) complex of pyridine-4- and pyrazine-2-hydroxamic acid as source of HNO: investigation of anti-tubercular and vasodilation activities.

A pharmacophore design approach, based on the coordination chemistry of an intimate molecular hybrid of active metabolites of pro-drugs, known to release active species upon enzymatic oxidative activation, is devised. This is exemplified by combining two anti-mycobacterial drugs: pyrazinamide (first line) and delamanid (third line) whose active metabolites are pyrazinoic acid (PyzCOOH) and likely nitroxyl (HNO (or NO.)), respectively. Aiming to generate those active species, a hybrid compound was envisaged by coordination of pyrazine-2-hydroxamic acid (PyzCONHOH) witha Na3[FeII(CN)5]moiety. The corresponding pentacyanoferrate(II) complex Na4[FeII(CN)5(PyzCONHO-)] was synthesized and characterized by several spectroscopic techniques, cyclic voltammetry, and DFT calculations. Chemical oxidation of this complex with H2O2 was shown to induce the release of the metabolite PyzCOOH, without the need of the Mycobacterium tuberculosis (Mtb) pyrazinamidase enzyme (PncA). Control experiments show that both H2O2- and N-coordinated pyrazine FeII species are required, ruling out a direct hydrolysis of the hydroxamic acid or an alternative oxidative route through chelation of ametal center by a hydroxamic group. The release of HNO was observed using EPR spectroscopy in the presence of a spin trapping agent. The devised iron metal complex of pyrazine-2-hydroxamic acid wasfound inactive against an actively growing/non-resistant Mtb strain; however, itshowed a strong dose-dependent and reversible vasodilatory activity with mostly lesser toxic effects than the reference drug sodium nitroprussiate, unveiling thus a potential indication for acute or chronic cardiovascular pathology. This is a priori a further indirect evidence of HNO release from this metal complex, standing as a possible pharmacophore model for an alternative vasodilator drug.

First unequivocal identification of the critical acyl radicals from the anti-tuberculosis drug isoniazid and its hydrazide analogs by complementary applications of ESR spin-trapping and HPLC/MS methods

The carbon-centered isonicotinic acyl radical of isoniazid (INH), a widely-used frontline anti-tuberculosis drug, has been considered to play a critical role in inhibiting Mycobacterium tuberculosis, but not fully identified. Here we show that this radical intermediate can be unequivocally characterized by complementary applications of ESR spin-trapping and HPLC/MS methods by employing N-tert-butyl-α-phenylnitrone (PBN) as the suitable spin-trapping agent, which can form the most stable radical adduct. More importantly, for the first time, analogous carbon-centered acyl radicals and their respective NAD+ adducts have also been detected and identified from its two isomers (nicotinic acid hydrazide and 2-pyridinecarbohydrazide) and benzhydrazide which are structurally-related to INH, but not by 2-chloroisonicotinohydrazide. This study represents the first unequivocal identification of the carbon-centered acyl radicals of INH and other hydrazide analogs by both ESR spin-trapping and HPLC/MS methods, which may have broad biomedical and toxicological significance for future research for more efficient hydrazide anti-tuberculosis drugs.

Potent Oxidation of DNA by Haloquinoid Disinfection Byproducts to the More Mutagenic Imidazolone dIz via an Unprecedented Haloquinone-Enoxy Radical-Mediated Mechanism.

Halogenated quinones are a class of carcinogenic intermediates and newly identified chlorination disinfection byproducts in drinking water. We found recently that halogenated quinones could enhance the decomposition of hydroperoxides independent of transition-metal ions and formation of the novel quinone enoxy/ketoxy radicals. Here, we show that the major oxidation product was 2-amino-5-[(2-deoxy-β-d-erythro-pentofuranosyl)amino]-4H-imidazol-4-one (dIz) when the nucleoside 2'-deoxyguanosine (dG) was treated with tetrachloro-1,4-benzoquinone (TCBQ) and t-butyl hydroperoxide (t-BuOOH). The formation of dIz was markedly inhibited by typical radical spin-trapping agents. Interestingly and unexpectedly, we found that the generated quinone enoxy radical played a critical role in dIz formation. Using [15N5]-8-oxodG, dIz was found to be produced either directly from dG or through the transient formation of 8-oxodG. Based on these data, we proposed that the production of dIz might be through an unusual haloquinone-enoxy radical-mediated mechanism. Analogous results were observed in the oxidation of ctDNA by TCBQ/t-BuOOH and when t-BuOOH was substituted by the endogenously generated physiologically relevant hydroperoxide 13S-hydroperoxy-9Z,11E-octadecadienoic acid. This is the first report that halogenated quinoid carcinogens and hydroperoxides can induce potent oxidation of dG to the more mutagenic product dIz via an unprecedented quinone-enoxy radical-mediated mechanism, which may partly explain their potential carcinogenicity.

An unexpected new pathway for nitroxide radical production via more reactve nitrogen-centered amidyl radical intermediate during detoxification of the carcinogenic halogenated quinones by N-alkyl hydroxamic acids

We found previously that nitroxide radical of desferrioxamine (DFO•) could be produced from the interaction between the classic iron chelating agent desferrioxamine (DFO, an N-alkyl trihydroxamic acid) and tetrachlorohydroquinone (TCHQ), one of the carconogenic quinoind metabolites of the widely used wood preservative pentachlorophenol. However, the underlying molecular mechanism remains unclear. Here N-methylacetohydroxamic acid (N-MeAHA) was synthesized and used as a simple model compound of DFO for further mechanistic study. As expected, direct ESR studies showed that nitroxide radical of N-MeAHA (Ac-(CH3)NO•) can be produced from N-MeAHA/TCHQ. Interestingly and unexpectedly, when TCHQ was substituted by its oxidation product tetrachloro-1,4-benzoquinone (TCBQ), although Ac-(CH3)NO• could also be produced, no concurrent formation of tetrachlorosemiquinone radical (TCSQ•) and TCHQ was detected, suggesting that Ac-(CH3)NO• did not result from direct oxidation of N-MeAHA by TCSQ• or TCBQ as proposed previously. To our surprise, a new nitrogen-centered amidyl radical was found to be generated from N-MeAHA/TCBQ, which was observed by ESR with the spin-trapping agents and further unequivacally identified as Ac-(CH3)N• by HPLC-MS. The final product of amidyl radical was isolated and identified as its corresponding amine. Analogous radical homolysis mechanism was observed with other halogenated quinoid compounds and N-alkyl hydroxamic acids including DFO. Interestingly, amidyl radicals were found to induce both DNA strand breaks and DNA adduct formation, suggesting that N-alkyl hydroxamic acids may exert their potential side-toxic effects via forming the reactive amidyl radical species. This study represents the first report of an unexpected new pathway for nitroxide radical production via hydrogen abstration reaction of a more reactive amidyl radical intermediate during the detoxification of the carcinogenic polyhalogenated quinones by N-alkyl hydroxamic acids, which provides more direct experimental evidence to better explain not only our previous finding that excess DFO can provide effective but only partial protection against TCHQ (or TCBQ)-induced biological damage, and also the potential side-toxic effects induced by DFO and other N-alkyl hydroxamic acid drugs.

The Study of the Neuroprotective Effect of the Extract from <i>Chelidonium Majus</i> L. <i>in Vitro</i>

The protection of neurons from damage and death is an important challenge in the development of treatment of brain ischemia and neurodegenerative diseases. This study aims to investigate protective effect of the extract prepared from Chelidonium majus, which contains total alkaloids. In the present study, we examined antioxidant activity of total alkaloids from Chelidonium majus in vitro. Hydroxyl radical and lipid radicals were detected using spin trapping agents with ESR spectrometer. Chelidonium majus extract exhibited dose-dependent scavenging effects on lipid radicals. Halfmaximal inhibitory concentration (IC50) of the extract was 2.96 mg/ml, whereas for hydroxyl radicals it was 55.13 mg/ ml. These results showed that extract of Chelidonium majus is partly inhibited free radicals. Antioxidant effects of this extract were further demonstrated by protecting enzyme activity of the mitochondrial respiratory electron transport chain (complex I) in isolated brain mitochondrial dysfunction induced by MDA. However, it did not change the decreased level of complex II, and malate dehydrogenase (MDH) in a concentration of 12 and 25 mg/ml. Here, we examined the neuroprotective effect of the extract from Chelidonium majus against oxidative stress in cultured cortical neurons, using MTT. We found that pretreatment with the extract of Chelidonium majus (0.05 and 0.5 mg/ml) significantly inhibited H2O2-induced cell death in cortical neurons.Furthermore, the use of a luciferase reporter (ARE-luc) assay showed that extract from Chelidonium majus activates protective signaling pathway in primary cortical neurons through ARE/Nrf2 pathway.Together, this suggests that total alkaloids from Chelidonium majus may be neuroprotective by increasing anti-oxidant gene expression.

Role of coke-bounded environmentally persistent free radicals in phenanthrene degradation by hydrogen peroxide

ABSTRACTEmission of polycyclic aromatic hydrocarbons (PAHs) is accompanied with the discharge of carbonaceous particles during the coke production. To degrade the adsorbed PAHs, hydrogen peroxide (H2O2) was applied as an oxidising agent, which might be activated by the inherent environmentally persistent free radicals (EPFRs) on coke particles. The transformation of phenanthrene (PHE), selected as model molecule, was achieved in H2O2/coke particle system without the addition of additional activating agent. This process consumed the particle-bounded EPFRs, inducing the decreasing of spin density from 1.92 × 1018 to 4.4 × 1017 spins g−1 in 30 min of reaction time. Electron paramagnetic resonance (EPR) technique coupled with spin-trapping agent 5, 5-dimethyl-1-pyrroline-N-oxide (DMPO) was used to probe the potential formation of reactive oxygen species. A higher capture [] concentration was observed with larger decreases in EPFRs concentration, indicating that EPFRs were the main contributor to the formation of . The obtained results suggested that the activation of H2O2 by EPFRs on coke particles resulted in the generation of hydroxyl radical (), which then back-reacted with adsorbed PHE. The finding of this study shed light on a new remediation technology for toxic carbonaceous byproducts discharged during the coke production.

Synthesis and characterization of Ce and Er doped ZrO2 nanoparticles as solar light driven photocatalysts

In this work, nanostructured rare-earth-doped zirconia was successfully synthesized, through a simple hydrothermal method, with a rare-earth oxide (REO)-based content varying from 0.5 to 5% molar (REO = CeO2; Er2O3). The samples were characterized by using several techniques, such as X-ray diffraction, transmission electron microscopy, X-ray photoelectron spectroscopy, diffuse reflectance spectroscopy. The photoactivity of the samples was tested under irradiation with simulated solar light by both the spin trapping technique, using DMPO (5,5-dimethyl-1-pyrroline-N-oxide) as spin trapping agent, and by the photodegradation of Rhodamine B, a common textile dye. All the doped samples resulted more active than the bare zirconia, and the correlation between the physical-chemical properties and the photoactivity of the materials has been investigated.

Improved Singlet Oxygen Detection Sensitivity in Electron Spin Resonance Using a Spin-trap Agent Incorporated into a Water-soluble Polymer Film

2,2,6,6-Tetramethyl-4-piperidinol (TEMP) is a spin-trap agent for electron spin resonance (ESR) that is usually employed in solution. However, in this study, TEMP is mixed with hydroxypropyl methyl...

Dose-dependent decrease in anti-oxidant capacity of whole blood after irradiation: A novel potential marker for biodosimetry

Many reports have demonstrated that radiation stimulates reactive oxygen species (ROS) production by mitochondria for a few hours to a few days after irradiation. However, these studies were performed using cell lines, and there is a lack of information about redox homeostasis in irradiated animals and humans. Blood redox homeostasis reflects the body condition well and can be used as a diagnostic marker. However, most redox homeostasis studies have focused on plasma or serum, and the anti-oxidant capacity of whole blood has scarcely been investigated. Here, we report changes in the anti-oxidant capacity of whole blood after X-ray irradiation using C57BL/6 J mice. Whole-blood anti-oxidant capacity was measured by electron spin resonance (ESR) spin trapping using a novel spin-trapping agent, 2-diphenylphosphinoyl-2-methyl-3,4-dihydro-2H-pyrrole N-oxide (DPhPMPO). We found that whole-blood anti-oxidant capacity decreased in a dose-dependent manner (correlation factor, r > 0.9; P < 0.05) from 2 to 24 days after irradiation with 0.5–3 Gy. We further found that the red blood cell (RBC) glutathione level decreased and lipid peroxidation level increased in a dose-dependent manner from 2 to 6 days after irradiation. These findings suggest that blood redox state may be a useful biomarker for estimating exposure doses during nuclear and/or radiation accidents.

Antiproliferative activity of novel isatinyl/indanyl nitrones (INs) as potential spin trapping agents of free radical intermediates.

A series of ketonitrones derived from isatin and indanone (INs) were synthesized and evaluated for their antiproliferative activities against several human cancer cell lines. Then, the antioxidant properties of these substrates were measured by the DPPH test to report their biological activity in terms of their spin trapping action. In particular, one substrate has showed very high biological and scavenging activity, probably due to the strong correlation between its spin trapping activity and structure.

Localized hydroxyl radical generation at mmol/L and mol/L levels in water by photon irradiation.

The generation of localized hydroxyl radical (•OH) in aqueous samples by low linear energy transfer irradiation was investigated. Several concentrations of 5,5-dimethyl-1-pyrroline-N-oxid solution (from 0.5 to 1,680 mmol/L) were prepared and irradiated with an identical dose of X-ray or γ-ray. The density of •OH generation in aqueous solution was evaluated by the electron paramagnetic resonance spin-trapping technique using 5,5-dimethyl-1-pyrroline-N-oxid as an electron paramagnetic resonance spin-trapping agent. The relationship between the molecular density of 5,5-dimethyl-1-pyrroline-N-oxid in the samples and the concentration of 5,5-dimethyl-1-pyrroline-N-oxid-OH generated in the irradiated samples was analyzed. Two different characteristic linear trends were observed in the 5,5-dimethyl-1-pyrroline-N-oxid-OH/5,5-dimethyl-1-pyrroline-N-oxid plots, which suggested •OH generation in two fashions, i.e., mmol/L- and mol/L-level local concentrations. The dose, dose rate, and/or the energy of photon irradiation did not affect the shapes of the 5,5-dimethyl-1-pyrroline-N-oxid-OH/5,5-dimethyl-1-pyrroline-N-oxid plots. Moreover, the addition of 5 mmol/L caffeine could cancel the contribution of mmol/L-level •OH generation, leaving a trace of mol/L-level •OH generation. Thus, the localized mmol/L- and mol/L-level generations of •OH, which were independent of experimental parameters such as dose, dose rate, and/or the energy of photon of low linear energy transfer radiation, were established.

In Vivo and In Situ Detection of Macromolecular Free Radicals Using Immuno-Spin Trapping and Molecular Magnetic Resonance Imaging.

In vivo free radical imaging in preclinical models of disease has become a reality. Free radicals have traditionally been characterized by electron spin resonance (ESR) or electron paramagnetic resonance (EPR) spectroscopy coupled with spin trapping. The disadvantage of the ESR/EPR approach is that spin adducts are short-lived due to biological reductive and/or oxidative processes. Immuno-spin trapping (IST) involves the use of an antibody that recognizes macromolecular 5,5-dimethyl-pyrroline-N-oxide (DMPO) spin adducts (anti-DMPO antibody), regardless of the oxidative/reductive state of trapped radical adducts. Recent Advances: The IST approach has been extended to an in vivo application that combines IST with molecular magnetic resonance imaging (mMRI). This combined IST-mMRI approach involves the use of a spin-trapping agent, DMPO, to trap free radicals in disease models, and administration of an mMRI probe, an anti-DMPO probe, which combines an antibody against DMPO-radical adducts and an MRI contrast agent, resulting in targeted free radical adduct detection. The combined IST-mMRI approach has been used in several rodent disease models, including diabetes, amyotrophic lateral sclerosis (ALS), gliomas, and septic encephalopathy. The advantage of this approach is that heterogeneous levels of trapped free radicals can be detected directly in vivo and in situ to pin point where free radicals are formed in different tissues. The approach can also be used to assess therapeutic agents that are either free radical scavengers or generate free radicals. Smaller probe constructs and radical identification approaches are being considered. The focus of this review is on the different applications that have been studied, advantages and limitations, and future directions. Antioxid. Redox Signal. 28, 1404-1415.