Nos tutelles

CNRS

Nos partenaires

Rechercher




Accueil

Productions scientifiques

Ouari, Olivier.

ACL - Articles dans des revues à comité de lecture

84 références.
2019

  • Besson, E.; Gastaldi, S.; Bloch, E.; Zielonka, J.; Zielonka, M.; Kalyanaraman, B.; Aslan, S.; Karoui, H.; Rockenbauer, A.; Ouari, O.; Hardy, M. Analyst 2019, 144 (14), 4194-4203.
    Résumé : The generation of superoxide radical anion in biological systems is one of the major initiating events in the redox biology of NADPH oxidases and mitochondrial redox signalling. However, the pallette of chemical tools for superoxide detection is very limited, hampering progress in understanding the chemical biology of superoxide. Although EPR spin trapping is regarded as the most rigorous technique for superoxide detection, rapid reduction of the EPR-active superoxide spin adducts to EPR-silent hydroxylamines, or to hydroxyl radical adducts by bioreductants, significantly limits the applicability of this technique in biological systems. To overcome these limitations, in this work, we report the synthesis and characterization of a new mesoporous silica functionalized with a phosphorylated cyclic spin trap (DIPPMPO nitrone). The DIPPMPO-grafted silica is a versatile spin-trap agent enabling the identification of a wide range of carbon or oxygen-centered transient radicals in organic and in aqueous media. Moreover, superoxide was efficiently trapped under in vitro conditions in both cell-free and cellular systems. The generated superoxide adduct exhibited an exceptional half-life of 3.5 h and a resistance toward bioreductant agents such as glutathione for several hours.

  • Bonucci, A.; Ouari, O.; Guigliarelli, B.; Belle, V.; Mileo, E. Chembiochem 2019.
    Résumé : Exploring the structure and dynamics of biomolecules in the context of their intracellular environment has become the ultimate challenge for structural biology. As the cellular environment is barely reproducible in vitro, investigation of biomolecules directly inside cells has attracted a growing interest. Among magnetic resonance approaches, site-directed spin labeling (SDSL) coupled to electron paramagnetic resonance (EPR) spectroscopy provides competitive and advantageous features to capture protein structure and dynamics inside cells. To date, several in-cell EPR approaches have been successfully applied to both bacterial and eukaryotic cells. In this review, the major advances of in-cell EPR spectroscopy are summarized, as well as the challenges this approach still poses.

  • Capozzi, A.; Patel, S.; Gunnarsson, C. P.; Marco-Rius, I.; Comment, A.; Karlsson, M.; Lerche, M. H.; Ouari, O.; Ardenkjaer-Larsen, J. H. Angewandte Chemie-International Edition 2019, 58 (5), 1334-1339.
    Résumé : Free radicals generated by UV-light irradiation of a frozen solution containing a fraction of pyruvic acid (PA) have demonstrated their dissolution dynamic nuclear polarization (dDNP) potential, providing up to 30% [1-C-13] PA liquid-state polarization. Moreover, their labile nature has proven to pave a way to nuclear polarization storage and transport. Herein, differently from the case of PA, the issue of providing dDNP UV-radical precursors (trimethylpyruvic acid and its methyl-deuterated form) not involved in any metabolic pathway was investigated. The C-13 dDNP performance was evaluated for hyperpolarization of [U-C-13(6),1,2,3,4,5,6,6-d(7)]-D-glucose. The generated UV-radicals proved to be versatile and highly efficient polarizing agents, providing, after dissolution and transfer (10 s), a C-13 liquid-state polarization of up to 32%.

  • Cheng, G.; Zhang, Q.; Pan, J.; Lee, Y.; Ouari, O.; Hardy, M.; Zielonka, M.; Myers, C. R.; Zielonka, J.; Weh, K.; Chang, A. C.; Chen, G.; Kresty, L.; Kalyanaraman, B.; You, M. Nature Communications 2019, 10, 2205.
    Résumé : Lung cancer often has a poor prognosis, with brain metastases a major reason for mortality. We modified lonidamine (LND), an antiglycolytic drug with limited efficacy, to mitochondria targeted mito-lonidamine (Mito-LND) which is 100-fold more potent. Mito-LND, a tumor selective inhibitor of oxidative phosphorylation, inhibits mitochondrial bioenergetics in lung cancer cells and mitigates lung cancer cell viability, growth, progression, and metastasis of lung cancer xenografts in mice. Mito-LND blocks lung tumor development and brain metastasis by inhibiting mitochondrial bioenergetics, stimulating the formation of reactive oxygen species, oxidizing mitochondrial peroxiredoxin, inactivating AKT/mTOR/p70S6K signaling, and inducing autophagic cell death in lung cancer cells. Mito-LND causes no toxicity in mice even when administered for eight weeks at 50 times the effective cancer inhibitory dose. Collectively, these findings show that mitochondrial targeting of LND is a promising therapeutic approach for investigating the role of autophagy in mitigating lung cancer development and brain metastasis.

  • Cheng, G.; Zielonka, J.; Hardy, M.; Ouari, O.; Chitambar, C. R.; Dwinell, M. B.; Kalyanaraman, B. Oncotarget 2019, 10 (37), 3518-3532.
    Résumé : We demonstrate that combined treatment with metformin (Met) or its mitochondria-targeted analog, mito-metformin (Mito-Met), and various iron chelators synergistically inhibits proliferation of pancreatic and triple-negative breast cancer cells. Previously, we reported that Met (at millimolar levels) and Mito-Met (at micromolar levels) inhibited pancreatic cancer cell proliferation. Inhibition of mitochondrial complex I and mitochondrial oxidative phosphorylation (OXPHOS) through stimulation of mitochondrial redox signaling was proposed as a key mechanism for decreased cancer cell proliferation. Because of its poor bioavailability, the use of Met as a "stand-alone" antitumor drug has been questioned. Iron chelators such as deferasirox (DFX) and dexrazoxane (DXR) exhibit antiproliferative effects in cancer cells. The potency of Met and Mito-Met was synergistically enhanced in the presence of iron chelators, DFX, N,N'-bis(2-hydroxyphenyl)ethylendiamine-N,N'-diacetic acid (HBED), and deferoxamine (DFO). Met, DXR (also ICRF-187), and DFO are FDA-approved drugs for treating diabetes, decreasing the incidence and severity of cardiotoxicity following chemotherapy, and mitigating iron toxicity, respectively. Thus, the synergistic antiproliferative effects of Met and Met analogs and iron chelators may have significant clinical relevance in cancer treatment. These findings may have implications in other OXPHOS-mediated cancer therapies.

  • Combes, S.; Tran, K. T.; Ayhan, M. M.; Karoui, H.; Rockenbauer, A.; Tonetto, A.; Monnier, V.; Charles, L.; Rosas, R.; Viel, S.; Siri, D.; Tordo, P.; Clair, S.; Wang, R.; Bardelang, D.; Ouari, O. Journal of the American Chemical Society 2019.
    Résumé : Triangular shapes have inspired scientists over time and are common in nature, such as the flower petals of oxalis triangularis, the triangular faces of tetrahedrite crystals, and the icosahedron faces of virus capsids. Supramolecular chemistry has enabled the construction of triangular assemblies, many of which possess functional features. Among these structures, cucurbiturils have been used to build supramolecular triangles, and we recently reported paramagnetic cucurbit[8]uril (CB[8]) triangles, but the reasons for their formation remain unclear. Several parameters have now been identified to explain their formation. At first sight, the radical nature of the guest was of prime importance in obtaining the triangles, and we focused on extending this concept to biradicals to get supramolecular hexaradicals. Two sodium ions were systematically observed by ESI-MS in trimer structures, and the presence of Na+ triggered or strengthened the triangulation of CB[8]/guest 1:1 complexes in solution. X-ray crystallography and molecular modeling have allowed the proposal of two plausible sites of residence for the two sodium cations. We then found that a diamagnetic guest with an H-bond acceptor function is equally good at forming CB[8] triangles. Hence, a guest molecule containing a ketone function has been precisely triangulated thanks to CB[8] and sodium cations as determined by DOSY-NMR and DLS. A binding constant for the triangulation of 1:1 to 3:3 complexes is proposed. This concept has finally been extended to the triangulation of ditopic guests toward network formation by the reticulation of CB[8] triangles using dinitroxide biradicals.

  • Salnikov, E. S.; Aussenac, F.; Abel, S.; Purea, A.; Tordo, P.; Ouari, O.; Bechinger, B. Solid State Nuclear Magnetic Resonance 2019, 100, 70-76.
    Résumé : Whereas specially designed dinitroxide biradicals, reconstitution protocols, oriented sample geometries and NMR probes have helped to much increase the DNP enhancement factors of membrane samples they still lag considerably behind those obtained from glasses made of protein solutions. Here we show that not only the MAS rotor material but also the distribution of the membrane samples within the NMR rotor have a pronounced effect on the DNP enhancement. These observations are rationalized with the cooling efficiency and the internal properties of the sample, monitored by their T-1 relaxation, microwave ON versus OFF signal intensities and DNP effect. The data are suggestive that for membranes the speed of cooling has a pronounced effect on the membrane properties and concomitantly the distribution of biradicals within the sample.

  • Viger-Gravel, J.; Avalos, C. E.; Kubicki, D. J.; Gajan, D.; Lelli, M.; Ouari, O.; Lesage, A.; Emsley, L. Angewandte Chemie-International Edition 2019, 58 (22), 7249-7253.
    Résumé : The introduction of high-frequency, high-power microwave sources, tailored biradicals, and low-temperature magic angle spinning (MAS) probes has led to a rapid development of hyperpolarization strategies for solids and frozen solutions, leading to large gains in NMR sensitivity. Here, we introduce a protocol for efficient hyperpolarization of F-19 nuclei in MAS DNP enhanced NMR spectroscopy. We identified trifluoroethanol-d(3) as a versatile glassy matrix and show that 12mm AMUPol (with microcrystalline KBr) provides direct F-19 DNP enhancements of over 100 at 9.4T. We applied this protocol to obtain DNP-enhanced F-19 and F-19-C-13 cross-polarization (CP) spectra for an active pharmaceutical ingredient and a fluorinated mesostructured hybrid material, using incipient wetness impregnation, with enhancements of approximately 25 and 10 in the bulk solid, respectively. This strategy is a general and straightforward method for obtaining enhanced F-19 MAS spectra from fluorinated materials.

  • Yang, X.; Giorgi, M.; Karoui, H.; Gigmes, D.; Hornebecq, V.; Ouari, O.; Kermagoret, A.; Bardelang, D. Chemical Communications 2019, 55 (92), 13824-13827.
    Résumé : Metal-organic framework (MOF) type crystals in which rigid, viologen-based pillars are surrounded by cucurbit[7]uril (CB[7]) macrocycles are described. These metal-organic rotaxane frameworks (MORF) are obtained by desolvation of a crystal precursor featuring a network already near-optimal toward 3D metal-ligand reticulation and show reversible photochromism.

  • Yin, H.; Cheng, Q.; Rosas, R.; Viel, S.; Monnier, V.; Charles, L.; Siri, D.; Gigmes, D.; Ouari, O.; Wang, R.; Kermagoret, A.; Bardelang, D. Chemistry-a European Journal 2019, 25 (54), 12552-12559.
    Résumé : The capture of pH-responsive guest molecules by CB[8] macrocycles made it possible to produce 2:2 host:guest complexes, releasing protons initially trapped on the guests, in a manner similar to the proton release observed in biology and operating through cascade supramolecular events. More information can be found in the Full Paper by R. Wang, A. Kermagoret, D. Bardelang, et al. (DOI: 10.1002/chem.201902057).
2018

  • Casano, G.; Karoui, H.; Ouari, O. Emagres 2018, 7 (4), 195-207.
    Résumé : In this article, we describe an in-depth overview of the development of paramagnetic polarizing agents for dynamic nuclear polarization (DNP)enhanced solid-state magic angle spinning (MAS) nuclear magnetic resonance (NMR). In DNP experiments, the large polarization of unpaired electrons is transferred to surrounding nuclei, which provides a maximum theoretical DNP enhancement of 660 for 1H NMR. The article includes a description of the different polarizing mechanisms and outlines key structural and magnetic parameters that contributed to the rational design of improved polarizing sources. The application of (di)nitroxides, heterobiradicals, narrow-line radicals, paramagnetic metal ions as well as site-specific polarizing agents is discussed. With the best polarizing agents, ssNMR MAS NMR/DNP enhances sensitivity by a factor of up to 200, providing decreased experiment time by five orders of magnitude and opening new avenues for NMR.

  • Cheng, G.; Zielonka, M.; Dranka, B.; Kumar, S. N.; Myers, C. R.; Bennett, B.; Garces, A. M.; Machado, L. G. D. D.; Thiebaut, D.; Ouari, O.; Hardy, M.; Zielonka, J.; Kalyanaraman, B. Journal of Biological Chemistry 2018, 293 (26), 10363-10380.
    Résumé : Reactive oxygen and nitrogen species (ROS/RNS) such as superoxide (O), hydrogen peroxide, lipid hydroperoxides, peroxynitrite, and hypochlorous and hypobromous acids play a key role in many pathophysiological processes. Recent studies have focused on mitochondrial ROS as redox signaling species responsible for promoting cell division, modulating and regulating kinases and phosphatases, and activating transcription factors. Many ROS also stimulate cell death and senescence. The extent to which these processes occur is attributed to ROS levels (low or high) in cells. However, the exact nature of ROS remains unknown. Investigators have used redox-active probes that, upon oxidation by ROS, yield products exhibiting fluorescence, chemiluminescence, or bioluminescence. Mitochondria-targeted probes can be used to detect ROS generated in mitochondria. However, because most of these redox-active probes (untargeted and mitochondria-targeted) are oxidized by several ROS species, attributing redox probe oxidation to specific ROS species is difficult. It is conceivable that redox-active probes are oxidized in common one-electron oxidation pathways, resulting in a radical intermediate that either reacts with another oxidant (including oxygen to produce O) and forms a stable fluorescent product or reacts with O to form a fluorescent marker product. Here, we propose the use of multiple probes and complementary techniques (HPLC, LC-MS, redox blotting, and EPR) and the measurement of intracellular probe uptake and specific marker products to identify specific ROS generated in cells. The low-temperature EPR technique developed to investigate cellular/mitochondrial oxidants can easily be extended to animal and human tissues.

  • Cheng, Q.; Yin, H.; Rosas, R.; Gigmes, D.; Ouari, O.; Wang, R.; Kermagoret, A.; Bardelang, D. Chemical Communications 2018, 54 (98), 13825-13828.
    Résumé : A molecular switch built with cucurbit[7]uril and a 3-station viologen-phenylene-imidazole compound exhibited pH actuated ring translocation with high fatigue resistance (up to 10(2) cycles). The switch movement was harnessed toward selectively masking the toxicity of the viologen fragment at neutral pH near non-cancerous cells, while exposing it at acid pH near cancer cells.

  • Hardy, M.; Zielonka, J.; Karoui, H.; Sikora, A.; Michalski, R.; Podsiadly, R.; Lopez, M.; Vasquez-Vivar, J.; Kalyanaraman, B.; Ouari, O. Antioxidants & Redox Signaling 2018.
    Résumé : Significance: Since the discovery of the superoxide dismutase enzyme, the generation and fate of short-lived oxidizing, nitrosating, nitrating, and halogenating species in biological systems has been of great interest. Despite the significance of reactive oxygen species (ROS) and reactive nitrogen species (RNS) in numerous diseases and intracellular signaling, the rigorous detection of ROS and RNS has remained a challenge. Recent Advances: Chemical characterization of the reactions of selected ROS and RNS with electron paramagnetic resonance (EPR) spin traps and fluorescent probes led to the establishment of species-specific products, which can be used for specific detection of several forms of ROS and RNS in cell-free systems and in cultured cells in vitro and in animals in vivo. Profiling oxidation products from the ROS and RNS probes provides a rigorous method for detection of those species in biological systems. Critical Issues: Formation and detection of species-specific products from the probes enables accurate characterization of the oxidative environment in cells. Measurement of the total signal (fluorescence, chemiluminescence, etc.) intensity does not allow for identification of the ROS/RNS formed. It is critical to identify the products formed by using chromatographic or other rigorous techniques. Product analyses should be accompanied by monitoring of the intracellular probe level, another factor controlling the yield of the product(s) formed. Future Directions: More work is required to characterize the chemical reactivity of the ROS/RNS probes, and to develop new probes/detection approaches enabling real-time, selective monitoring of the specific products formed from the probes.
    Mots-clés : amplex red assay, boronate probes, diagnostic marker products, fluorescent-probes, hydroethidine, hydrogen-peroxide, hypochlorous acid, in-vivo, living cells, nitric-oxide synthase, nitronyl nitroxides, nitronyl nitroxides, peroxynitrite, spin trapping, spin-trapping properties, spin-trapping properties, superoxide radical anion.

  • Jaudzems, K.; Bertarello, A.; Chaudhari, S. R.; Pica, A.; Cala-De Paepe, D.; Barbet-Massin, E.; Pell, A. J.; Akopjana, I.; Kotelovica, S.; Gajan, D.; Ouari, O.; Tars, K.; Pintacuda, G.; Lesage, A. Angewandte Chemie-International Edition 2018, 57 (25), 7458-7462.
    Résumé : Dynamic nuclear polarization (DNP) is a powerful way to overcome the sensitivity limitation of magic-angle-spinning (MAS) NMR experiments. However, the resolution of the DNPNMR spectra of proteins is compromised by severe line broadening associated with the necessity to perform experiments at cryogenic temperatures and in the presence of paramagnetic radicals. High-quality DNP-enhanced NMR spectra of the Acinetobacter phage 205 (AP205) nucleocapsid can be obtained by combining high magnetic field (800MHz) and fast MAS (40kHz). These conditions yield enhanced resolution and long coherence lifetimes allowing the acquisition of resolved 2D correlation spectra and of previously unfeasible scalar-based experiments. This enables the assignment of aromatic resonances of the AP205 coat protein and its packaged RNA, as well as the detection of long-range contacts, which are not observed at room temperature, opening new possibilities for structure determination.

  • Kalyanaraman, B.; Chen, G.; Hardy, M.; Ouari, O.; Lopez, M.; Joseph, J.; Zielonka, J.; Dwinell, M. B. Redox Biology 2018, 16, 426-427.

  • Kalyanaraman, B.; Cheng, G.; Hardy, M.; Ouari, O.; Bennett, B.; Zielonka, J. Redox Biology 2018, 15, 347-362.
    Résumé : Reactive oxygen species (ROS) have been implicated in tumorigenesis (tumor initiation, tumor progression, and metastasis). Of the many cellular sources of ROS generation, the mitochondria and the NADPH oxidase family of enzymes are possibly the most prevalent intracellular sources. In this article, we discuss the methodologies to detect mitochondria-derived superoxide and hydrogen peroxide using conventional probes as well as newly developed assays and probes, and the necessity of characterizing the diagnostic marker products with HPLC and LC-MS in order to rigorously identify the oxidizing species. The redox signaling roles of mitochondrial ROS, mitochondrial thiol peroxidases, and transcription factors in response to mitochondria-targeted drugs are highlighted. ROS generation and ROS detoxification in drug-resistant cancer cells and the relationship to metabolic reprogramming are discussed. Understanding the subtle role of ROS in redox signaling and in tumor proliferation, progression, and metastasis as well as the molecular and cellular mechanisms (e.g., autophagy) could help in the development of combination therapies. The paradoxical aspects of antioxidants in cancer treatment are highlighted in relation to the ROS mechanisms in normal and cancer cells. Finally, the potential uses of newly synthesized exomarker probes for in vivo superoxide and hydrogen peroxide detection and the low-temperature electron paramagnetic resonance technique for monitoring oxidant production in tumor tissues are discussed.

  • Kalyanaraman, B.; Cheng, G.; Hardy, M.; Ouari, O.; Lopez, M.; Joseph, J.; Zielonka, J.; Dwinell, M. B. Redox Biology 2018, 14, 316-327.
    Résumé : The present review is a sequel to the previous review on cancer metabolism published in this journal. This review focuses on the selective antiproliferative and cytotoxic effects of mitochondria-targeted therapeutics (MTTs) in cancer cells. Emerging research reveals a key role of mitochondrial respiration on tumor proliferation. Previously, a mitochondria-targeted nitroxide was shown to selectively inhibit colon cancer cell proliferation at submicromolar levels. This review is centered on the therapeutic use of MTTs and their bioenergetic profiling in cancer cells. Triphenylphosphonium cation conjugated to a parent molecule (e.g., vitamin-E or chromanol, ubiquinone, and metformin) via a linker alkyl chain is considered an MTT. MTTs selectively and potently inhibit proliferation of cancer cells and, in some cases, induce cytotoxicity. MTTs inhibit mitochondrial complex I activity and induce mitochondrial stress in cancer cells through generation of reactive oxygen species. MTTs in combination with glycolytic inhibitors synergistically inhibit tumor cell proliferation. This review discusses how signaling molecules traditionally linked to tumor cell proliferation affect tumor metabolism and bioenergetics (glycolysis, TCA cycle, and glutaminolysis).

  • Karthikeyan, G.; Bonucci, A.; Casano, G.; Gerbaud, G.; Abel, S.; Thome, V.; Kodjabachian, L.; Magalon, A.; Guigliarelli, B.; Belle, V.; Ouari, O.; Mileo, E. Angewandte Chemie-International Edition 2018, 57 (5), 1366-1370.
    Résumé : Approaching protein structural dynamics and protein-protein interactions in the cellular environment is a fundamental challenge. Owing to its absolute sensitivity and to its selectivity to paramagnetic species, site-directed spin labeling (SDSL) combined with electron paramagnetic resonance (EPR) has the potential to evolve into an efficient method to follow conformational changes in proteins directly inside cells. Until now, the use of nitroxide-based spin labels for in-cell studies has represented a major hurdle because of their short persistence in the cellular context. The design and synthesis of the first maleimido-proxyl-based spin label (M-TETPO) resistant towards reduction and being efficient to probe protein dynamics by continuous wave and pulsed EPR is presented. In particular, the extended lifetime of M-TETPO enabled the study of structural features of a chaperone in the absence and presence of its binding partner at endogenous concentration directly inside cells.

  • Marco-Rius, I.; Cheng, T.; Gaunt, A. P.; Patel, S.; Kreis, F.; Capozzi, A.; Wright, A. J.; Brindle, K. M.; Ouari, O.; Comment, A. Journal of the American Chemical Society 2018, 140 (43), 14455-14463.
    Résumé : Whether for C-13 magnetic resonance studies in chemistry, biochemistry, or biomedicine, hyperpolarization methods based on dynamic nuclear polarization (DNP) have become ubiquitous. DNP requires a source of unpaired electrons, which are commonly added to the sample to be hyperpolarized in the form of stable free radicals. Once polarized, the presence of these radicals is unwanted. These radicals can be replaced by nonpersistent radicals created by the photoirradiation of pyruvic acid (PA), which are annihilated upon dissolution or thermalization in the solid state. However, since PA is readily metabolized by most cells, its presence may be undesirable for some metabolic studies. In addition, some C-13 substrates are photosensitive and therefore may degrade during the photogeneration of a PA radical, which requires ultraviolet (UV) light. We show here that the photoirradiation of phenylglyoxylic acid (PhGA) using visible light produces a nonpersistent radical that, in principle, can be used to hyperpolarize any molecule. We compare radical yields in samples containing PA and PhGA upon photoirradiation with broadband and narrowband UV-visible light sources. To demonstrate the suitability of PhGA as a radical precursor for DNP, we polarized the gluconeogenic probe C-13-dihydroxyacetone, which is UV-sensitive, using a commercial 3.35 T DNP polarizer and then injected this into a mouse and followed its metabolism in vivo.

  • Ouari, O.; Bardelang, D. Israel Journal of Chemistry 2018, 58 (3-4), 343-356.
    Résumé : The interface of nitroxide radicals with cucurbit[n]urils (CB[n]) is covered in this review. Either used as spin probes, spin labels, or stoppers for paramagnetic rotaxanes, nitroxides offer new opportunities to investigate CB[n] and their inclusion complexes, especially thanks to EPR spectroscopy. On the other hand, CB[n] also offer interesting opportunities to modulate nitroxide properties such as protection or structuring. The multiple back and forth between the two well-established areas where nitroxides bring information about CB[n], and CB[n] modulate nitroxide properties, have enabled to shed some light at this new interesting interface.

  • Ouedrago, M.; Abou, A.; Djande, A.; Ouari, O.; Zoueu, T. J. Acta Crystallographica Section E-Crystallographic Communications 2018, 74, 530-+.
    Résumé : In the title compound, C20H18O4, the benzoate ring is oriented at an acute angle of 33.10 (12)degrees with respect to the planar (r.m.s deviation = 0.016 angstrom) coumarin ring system. An intramolecular C-H center dot center dot center dot O hydrogen bond closes an S(6) ring motif. In the crystal, C-H center dot center dot center dot O contacts generate infinite C(6) chains along the b-axis direction. Also present are pi-pi stacking interactions between neighbouring pyrone and benzene rings [centroid-centroid distance = 3.7034 (18) A degrees] and C=O center dot center dot center dot pi interactions [O center dot center dot center dot centroid = 3.760 (3) angstrom]. The data obtained from quantum chemical calculations performed on the title compound are in good agreement with the observed structure, although the calculated C-O-C-C torsion angle between the coumarin ring system and the benzoate ring (129.1 degrees) is somewhat lower than the observed value [141.3 (3)degrees]. Hirshfeld surface analysis has been used to confirm and quantify the supramolecular interactions.

  • Wisser, D.; Karthikeyan, G.; Lund, A.; Casano, G.; Karoui, H.; Yulikov, M.; Menzildjian, G.; Pinon, A. C.; Purea, A.; Engelke, F.; Chaudhari, S. R.; Kubicki, D.; Rossini, A. J.; Moroz, I. B.; Gajan, D.; Coperet, C.; Jeschke, G.; Lelli, M.; Emsley, L.; Lesage, A.; Ouari, O. Journal of the American Chemical Society 2018, 140 (41), 13340-13349.
    Résumé : Dynamic nuclear polarization (DNP) solid-state nuclear magnetic resonance (NMR) has developed into an invaluable tool for the investigation of a wide range of materials. However, the sensitivity gain achieved with many polarizing agents suffers from an unfavorable field and magic angle spinning (MAS) frequency dependence. We present a series of new hybrid biradicals, soluble in organic solvents, that consist of an isotropic narrow electron paramagnetic resonance line radical, alpha,gamma-isdiphenylene-beta-phenylallyl (BDPA), tethered to a broad line nitroxide. By tuning the distance between the two electrons and the substituents at the nitroxide moiety, correlations between the electron-electron interactions and the electron spin relaxation times on one hand and the DNP enhancement factors on the other hand are established. The best radical in this series has a short methylene linker and bears bulky phenyl spirocyclohexyl ligands. In a 1.3 mm prototype DNP probe, it yields enhancements of up to 185 at 18.8 T (800 MHz H-1 resonance frequency) and 40 kHz MAS. We show that this radical gives enhancement factors of over 60 in 3.2 mm sapphire rotors at both 18.8 and 21.1 T (900 MHz H-1 resonance frequency), the highest magnetic field available today for DNP. The effect of the rotor size and of the microwave irradiation inside the MAS rotor is discussed. Finally, we demonstrate the potential of this new series of polarizing agents by recording high field Al-27 and Si-29 DNP surface enhanced NMR spectra of amorphous aluminosilicates and O-17 NMR on silica nanoparticles.

  • Zhao, X.; Liu, F.; Zhao, Z.; Karoui, H.; Bardelang, D.; Ouari, O.; Liu, S. Organic & Biomolecular Chemistry 2018, 16 (20), 3809-3815.
    Résumé : Most 1,4,5,8-naphthalenediimide (NDI) derivatives, especially those with mild -acidity cannot achieve photoinduced electron-transfer (PET). Here we report our investigations on the binding interactions of a NDI diammonium dichloride salt (NDICl2) with cucurbit[n]uril (CB[n], n = 8, 10) and the formation process of a NDI radical anion upon photoexcitation of the NDI derivative in the presence of CB[n] (n = 8, 10). As a comparison, the influence of CB[7] on the PET process was also evaluated. The results show that the NDI core can be partially or fully encapsulated in CB[8] or CB[10] to form inclusion complexes at molar ratios of 1:1 (CB[8]NDI2+) or 1:2 (CB[10]2NDI(2+)). In the presence of the host, quick formation of the NDI radical anion was observed with respect to no radical anion formation without the host. According to the spectral results, interesting CB[8]-promoted charge-transfer interactions between the NDI radical anion and NDI2+ are assumed during
2017

  • Chaudhari, S. R.; Wisser, D.; Pinon, A. C.; Berruyer, P.; Gajan, D.; Tordo, P.; Ouari, O.; Reiter, C.; Engelke, F.; Coperet, C.; Lelli, M.; Lesage, A.; Emsley, L. Journal of the American Chemical Society 2017, 139 (31), 10609–10612.
    Résumé : Dynamic nuclear polarization (DNP) has recently emerged as a tool to enhance the sensitivity of solid-state NMR experiments. However, so far high enhancements (\textgreater100) are limited to relatively low magnetic fields, and DNP at fields higher than 9.4 T significantly drops in efficiency. Here we report solid-state Overhauser effect DNP enhancements of over 100 at 18.8 T. This is achieved through the unexpected discovery that enhancements increase rapidly with increasing magic angle spinning (MAS) rates. The measurements are made using 1,3-bisdiphenylene-2-phenylallyl dissolved in o-terphenyl at 40 kHz MAS. We introduce a source sink diffusion model for polarization transfer which is capable of explaining the experimental observations. The advantage of this approach is demonstrated on mesoporous alumina with the acquisition of well-resolved DNP surface enhanced Al-27 cross-polarization spectra.
    Mots-clés : enhanced nmr-spectroscopy, high-frequency, mas-dnp, polarizing agents, rotating solids, sensitivity, sites, solid-state nmr, surface, temperature.

  • Kalyanaraman, B.; Cheng, G.; Hardy, M.; Ouari, O.; Sikora, A.; Zielonka, J.; Dwinell, M. Interface Focus 2017, 7 (2), 20160109.
    Résumé : Reports suggest that metformin exerts anti-cancer effects in diabetic individuals with pancreatic cancer. Thus, metformin is currently being repurposed as a potential drug in cancer treatment. Studies indicate that potent metformin analogues are required in cancer treatment because of the low bioavailability of metformin in humans at conventional antidiabetic doses. We proposed that improved mitochondrial targeting of metformin by attaching a positively charged lipophilic triphenylphosphonium group will result in a new class of mitochondria-targeted metformin analogues with significantly enhanced anti-tumour potential. Using this approach, we synthesized various mitochondria-targeted metformin analogues with different alkyl chain lengths. Results indicate that the antiproliferative effects increased with increasing alkyl chain lengths (100-fold to 1000-fold). The lead compound, mito-metformin(10), potently inhibited mitochondrial respiration through inhibition of complex I, stimulation of superoxide and hydrogen peroxide formation and activation of AMPK. When used in combination with ionizing radiation, mito-metformin(10) acted as a radiosensitizer of pancreatic cancer cells. Because of the 1000-fold-higher potency of mitochondria-targeted metformin(10), therapeutically effective plasma concentrations likely can be achieved in cancer patients.
    Mots-clés : ampk, analogs, anti-cancer agent, biguanides, bioenergetics, cancer-cell sensitivity, drugs, endothelium, generation, metabolism, metformin, mitochondria, peroxynitrite, radiosensitization, Reactive oxygen species, redox signalling.

  • Kalyanaraman, B.; Cheng, G.; Hardy, M.; Ouari, O.; Sikora, A.; Zielonka, J.; Dwinell, M. B. Cell Biochemistry and Biophysics 2017, 75 (3-4), 311–317.
    Résumé : Metformin, one of the most widely prescribed antidiabetic drugs in the world, is being repurposed as a potential drug in cancer treatment. Epidemiological studies suggest that metformin exerts anticancer effects in diabetic patients with pancreatic cancer. However, at typical antidiabetic doses the bioavailability of metformin is presumably too low to exert antitumor effects. Thus, more potent analogs of metformin are needed in order to increase its anticancer efficacy. To this end, a new class of mitochondria-targeted metformin analogs (or mito-metformins) containing a positively-charged lipophilic triphenylphosphonium group was synthesized and tested for their antitumor efficacy in pancreatic cancer cells. Results indicate that the lead compound, mito-metformin(10), was nearly 1000-fold more potent than metformin in inhibiting mitochondrial complex I activity, inducing reactive oxygen species (superoxide and hydrogen peroxide) that stimulate redox signaling mechanisms, including the activation of adenosinemonophosphate kinase and inhibition of proliferation of pancreatic cancer cells. The potential use of the low-temperature electron paramagnetic resonance technique in assessing the role of mitochondrial complexes including complex I in tumor regression in response to metformin and mito-metformins in the in vivo setting is discussed.
    Mots-clés : analogs, biguanides, bioenergetics, cell-death, energy-metabolism, hydroethidine, identification, intracellular superoxide, Mitochondrial metabolism, Modified metformin, Pancreatic cancer, pancreatic-cancer, peroxynitrite, Reactive oxygen species, Redox signaling.

  • Lescic, S.; Karoui, H.; Hardy, M.; Charles, L.; Tordo, P.; Ouari, O.; Gaudel-Siri, A.; Siri, D. Journal of Physical Organic Chemistry 2017, 30 (10), e3677.
    Résumé : Spin trapping coupled with electron paramagnetic resonance spectroscopy is the most direct method for detection of very low concentrations of free radicals, and it has been intensively used to detect superoxide or alkylperoxyl radicals in biological systems, using cyclic nitrones as spin traps. The half-life time of the ensuing spin adducts depends dramatically on the spin trap structure; however, their mechanism of decay has never been definitely established. We investigated at the MRMP2/CASSCF (MultiReference second-order Moller-Plesset perturbation theory/Complete Active Set Self Consistent Field) level of theory the mechanism of decay of methylperoxyl and tert-butylperoxyl spin adducts formed with various cyclic nitrones. We showed that no transition state can be located for the O-O homolytic cleavage, which yields an intermediate biradical with the following sequence O-center dot-N-C-O-center dot. Then, homolytic cleavage of the N-C bond yields a nitrosoaldehyde, through an early transition state with a very low activation energy. For each nitrone used as spin trap, electrospray ionization mass spectrometry analysis of the spin trapping mixture allowed to detect the presence of the expected nitrosoaldehyde. We generated tert-butylperoxyl spin adducts in toluene, and we found a good correlation between their half-life time and the bond dissociation energy of their peroxidic O-O bond. The theoretical method was then extended to hydroperoxyl spin adducts in water and provided promising results.
    Mots-clés : 2nd-order perturbation-theory, alkylperoxyl spin adducts, biradical, centered radicals, cyclic nitrones, depmpo, dft, dmpo, epr, half-life time, mrmp2/casscf, radical-anion adduct, spin trapping, superoxide, trapping properties.

  • Liao, W. - C.; Ong, T. - C.; Gajan, D.; Bernada, F.; Sauvee, C.; Yulikov, M.; Pucino, M.; Schowner, R.; Schwarzwalder, M.; Buchmeiser, M. R.; Jeschke, G.; Tordo, P.; Ouari, O.; Lesage, A.; Emsley, L.; Coperet, C. Chemical Science 2017, 8 (1), 416–422.
    Résumé : Dynamic Nuclear Polarization Surface Enhanced NMR Spectroscopy (DNP SENS) is an effective method to significantly improve solid-state NMR investigation of solid surfaces. The presence of unpaired electrons (polarizing agents) is crucial for DNP, but it has drawbacks such as leading to faster nuclear spin relaxation, or even reaction with the substrate under investigation. The latter can be a particular problem for heterogeneous catalysts. Here, we present a series of carbosilane-based dendritic polarizing agents, in which the bulky dendrimer can reduce the interaction between the solid surface and the free radical. We thereby preserve long nuclear T-2 of the surface species, and even successfully enhance a reactive heterogeneous metathesis catalyst.
    Mots-clés : biradicals, chemistry, complexes, dynamic nuclear-polarization, efficient, enhanced nmr-spectroscopy, metathesis catalysts, sites, sn-beta zeolite, tempo.

  • Poulhes, F.; Rizzato, E.; Bernasconi, P.; Rosas, R.; Viel, S.; Jicsinszky, L.; Rockenbauer, A.; Bardelang, D.; Siri, D.; Gaudel-Siri, A.; Karoui, H.; Hardy, M.; Ouari, O. Organic & Biomolecular Chemistry 2017, 15 (30), 6358–6366.
    Résumé : Three new DEPMPO-based spin traps have been designed and synthesized for improved superoxide detection, each carrying a cyclodextrin (CD) moiety but with a different alkyl chain on the phosphorus atom or with a long spacer arm. EPR spectroscopy allowed us to estimate the half-life of the superoxide spin adducts which is close to the value previously reported for CD-DEPMPO (t(1/2) approximate to 50-55 min under the conditions investigated). The spectra are typical of superoxide adducts (almost no features of the HO center dot adduct that usually forms with time for other nitrone spin traps such as DMPO) and we show that at 250 mu M, the new spin trap enables the reliable detection of superoxide by 1 scan at the position opposite to the corresponding spin trap without the CD moiety. The resistance of the spin adducts to a reduction process has been evaluated, and the superoxide spin adducts are sensitive to ascorbate and glutathione (GSH), but not to glutathione peroxidase/GSH, reflecting the exposed nature of the nitroxide moiety to the bulk solvent. To understand these results, 2D-ROESY NMR studies and molecular dynamics pointed to a shallow or surface self-inclusion of the nitrone spin traps and of nitroxide spin adducts presumably due to the high flexibility of the permethyl-beta-CD rim.
    Mots-clés : conjugate, cyclic nitrone, depmpo, epr, in-vivo, inclusion, metabolic stability, radical adducts, resonance, trapping properties.

  • Sikora, A.; Michalski, R.; Hardy, M.; Ouari, O.; Adamus, J.; Marcinek, A.; Zielonka, J.; Kalyanaraman, B. Free Radical Biology and Medicine 2017, 108, S38–S38.
    Mots-clés : hydroethidine, superoxide radical anion.

  • Viger-Gravel, J.; Berruyer, P.; Gajan, D.; Basset, J. - M.; Lesage, A.; Tordo, P.; Ouari, O.; Emsley, L. Angewandte Chemie-International Edition 2017, 56 (30), 8726–8730.
    Résumé : Aqueous acrylamide gels can be used to provide dynamic nuclear polarization (DNP) NMR signal enhancements of around 200 at 9.4 T and 100 K. The enhancements are shown to increase with crosslinker concentration and low concentrations of the AMUPol biradical. This DNP matrix can be used in situations where conventional incipient wetness methods fail, such as to obtain DNP surface enhanced NMR spectra from inorganic nanoparticles. In particular, we obtain Cd-113 spectra from CdTe-COOH NPs in minutes. The spectra clearly indicate a highly disordered cadmium-rich surface.
    Mots-clés : acrylamide gels, angle-spinning nmr, correlation spectroscopy, dynamic nuclear polarization, high-frequency, nanoparticles, performance, polarizing agents, polyacrylamide gel, polymer gels, proteins, quantum dots, sensitivity, solid-state nmr.

  • Yin, H.; Dumur, F.; Niu, Y.; Ayhan, M. M.; Grauby, O.; Liu, W.; Wang, C.; Siri, D.; Rosas, R.; Tonetto, A.; Gigmes, D.; Wang, R.; Bardelang, D.; Ouari, O. Acs Applied Materials & Interfaces 2017, 9 (38), 33220–33228.
    Résumé : This work describes latent fluorescence particles (LFPs) based on a new environmentally sensitive carbazole compound aggregated in water and their use as sensors for probing various cavitands and the different stages of aggregating systems. Cyclodextrins (CDs), cucurbit[n]urils (CB[n], n = 6, 7,'8), and a resorcinarene capsule were used to study the dynamic nature of the LFPs. The fluorescence was dramatically enhanced by a proposed disaggregation-induced emission enhancement (DIEE) mechanism with specific features for CB [n]. Then, the aggregated states of the dipeptides Leu-Leu, Phe-Phe, and Fmoc-Leu-Leu (vesicles, crystals, fibers) were studied by fluorescence spectroscopy and confocal fluorescence microscopy thanks to the adaptive and emissive behavior of the LFPs, allowing us to study an interesting polymorphism phenomenon. The LFPs have then been used in the sensing of the aggregation of the polysaccharide alginate, for which distinct fluorescence turn-on is detected upon stepwise biopolymer assembly, and for amylose detection. The carbazole particles not only adapt to various environments but also display multicolor fluorescent signals. They can be used for the fast probing of the aggregation propensity of newly prepared molecules or biologically relevant compounds or to accelerate the discovery of new macrocycles or of self-assembling peptides in water.
    Mots-clés : assemblies, binding, chameleonic dye, cucurbit[n]urils, disaggregation-induced emission enhancement, emission, fluorescent dyes, fluorophore, host-guest complexes, latent fluorescence particles, light, nanostructures, peptide aggregation, polysaccharide aggregation, quantum dots, water.

  • Zielonka, J.; Joseph, J.; Sikora, A.; Hardy, M.; Ouari, O.; Vasquez-Vivar, J.; Cheng, G.; Lopez, M.; Kalyanaraman, B. Chemical Reviews 2017, 117 (15), 10043–10120.
    Résumé : Mitochondria are recognized as one of the most important targets for new drug design in cancer, cardiovascular, and neurological diseases. Currently, the most effective way to deliver drugs specifically to mitochondria is by covalent linking a lipophilic cation such as an alkyltriphenylphosphonium moiety to a pharmacophore of interest. Other delocalized lipophilic cations, such as rhodamine, natural and synthetic mitochondria-targeting peptides, and nanoparticle vehicles, have, also been used for mitochondrial delivery of small molecules. Depending on the approach used, and the cell and mitochondrial membrane potentials, more than 1000-fold higher mitochondrial concentration can be achieved. Mitochondrial targeting has been developed to study mitochondrial physiology and dysfunction and the interaction between mitochondria and other subcellular organelles and for treatment of a variety of diseases such as neurodegeneration and cancer. In this Review, we discuss efforts to target small-molecule compounds to mitochondria for probing mitochondria function, as. diagnostic tools and potential therapeutics. We describe the physicochemical basis for mitochondrial accumulation of lipophilic cations, synthetic chemistry strategies to target compounds to mitochondria, mitochondrial probes, and sensors, and examples of mitochondrial targeting of bioactive compounds. Finally, we review published attempts to apply mitochondria-targeted agents for the treatment of cancer and neurodegenerative diseases.
    Mots-clés : aggregation-induced-emission, alpha-tocopheryl succinate, biomembrane-produced energy, cells-in-vitro, hydrogen-sulfide donor, ischemia-reperfusion injury, mass-spectrometry probe, pancreatic-cancer cells, positron-emission-tomography, ratiometric fluorescent-probe.
2016

  • Benchekroun, M.; Romero, A.; Egea, J.; Leon, R.; Michalska, P.; Buendia, I.; Jimeno, M. L.; Jun, D.; Janockova, J.; Sepsova, V.; Soukup, O.; Bautista-Aguilera, O. M.; Refouvelet, B.; Ouari, O.; Marco-Contelles, J.; Ismaili, L. Journal of Medicinal Chemistry 2016, 59 (21), 9967-9973.
    Résumé : Novel multifunctional tacrines for Alzheimer's disease were obtained by Ugi-reaction between ferulic (or lipoic acid), a melatonin-like isocyanide, formaldehyde, and tacrine derivatives, according to the antioxidant additive approach in order to modulate the oxidative stress as therapeutic strategy. Compound 5c has been identified as a promising permeable agent showing excellent antioxidant properties, strong cholinesterase inhibitory activity, less hepatotoxicity than tacrine, and the best neuroprotective capacity, being able to significantly activate the Nrf2 transcriptional pathway.
    Mots-clés : acetylcholinesterase, ache inhibitors, derivatives, design, drug candidates, heterodimers, hybrids, hypothesis, mechanisms, neurodegenerative diseases.

  • Cheng, G.; Zielonka, J.; Ouari, O.; Lopez, M.; McAllister, D.; Boyle, K.; Barrios, C. S.; Weber, J. J.; Johnson, B. D.; Hardy, M.; Dwinell, M. B.; Kalyanaraman, B. Cancer Research 2016, 76 (13), 3904-3915.
    Résumé : Metformin (Met) is an approved antidiabetic drug currently being explored for repurposing in cancer treatment based on recent evidence of its apparent chemopreventive properties. Met is weakly cationic and targets the mitochondria to induce cytotoxic effects in tumor cells, albeit not very effectively. We hypothesized that increasing its mitochondria-targeting potential by attaching a positively charged lipophilic substituent would enhance the antitumor activity of Met. In pursuit of this question, we synthesized a set of mitochondria-targeted Met analogues (Mito-Mets) with varying alkyl chain lengths containing a triphenylphosphonium cation (TPP+). In particular, the analogue Mito-Met(10), synthesized by attaching TPP+ to Met via a 10-carbon aliphatic side chain, was nearly 1,000 times more efficacious than Met at inhibiting cell proliferation in pancreatic ductal adenocarcinoma (PDAC). Notably, in PDAC cells, Mito-Met(10) potently inhibited mitochondrial complex I, stimulating superoxide and AMPK activation, but had no effect in nontransformed control cells. Moreover, Mito-Met(10) potently triggered G(1) cell-cycle phase arrest in PDAC cells, enhanced their radiosensitivity, and more potently abrogated PDAC growth in preclinical mouse models, compared with Met. Collectively, our findings show how improving the mitochondrial targeting of Met enhances its anticancer activities, including aggressive cancers like PDAC in great need of more effective therapeutic options. (C) 2016 AACR.
    Mots-clés : bioenergetics, breast-cancer, death, generation, ionizing-radiation, kinase ampk, lung-cancer, metabolism, proliferation, survival.

  • Kubicki, D. J.; Casano, G.; Schwarzwaelder, M.; Abel, S.; Sauvee, C.; Ganesan, K.; Yulikov, M.; Rossini, A. J.; Jeschke, G.; Coperet, C.; Lesage, A.; Tordo, P.; Ouari, O.; Emsley, L. Chemical Science 2016, 7 (1), 550-558.
    Résumé : A series of 37 dinitroxide biradicals have been prepared and their performance studied as polarizing agents in cross-effect DNP NMR experiments at 9.4 T and 100 K in 1,1,2,2-tetrachloroethane (TCE). We observe that in this regime the DNP performance is strongly correlated with the substituents on the polarizing agents, and electron and nuclear spin relaxation times, with longer relaxation times leading to better enhancements. We also observe that deuteration of the radicals generally leads to better DNP enhancement but with longer build-up time. One of the new radicals introduced here provides the best performance obtained so far under these conditions.
    Mots-clés : c-13, dnp, enhanced nmr, nanoparticles, polarizing agents, resonance, signal enhancement, solid-state nmr, spectroscopy, temperatures.

  • Salnikov, E. S.; Aisenbrey, C.; Aussenac, F.; Ouari, O.; Sarrouj, H.; Reiter, C.; Tordo, P.; Engelke, F.; Bechinger, B. Scientific Reports 2016, 6, 20895.
    Résumé : Dynamic Nuclear Polarization (DNP) has been introduced to overcome the sensitivity limitations of nuclear magnetic resonance (NMR) spectroscopy also of supported lipid bilayers. When investigated by solid-state NMR techniques the approach typically involves doping the samples with biradicals and their investigation at cryo-temperatures. Here we investigated the effects of temperature and membrane hydration on the topology of amphipathic and hydrophobic membrane polypeptides. Although the antimicrobial PGLa peptide in dimyristoyl phospholipids is particularly sensitive to topological alterations, the DNP conditions represent well its membrane alignment also found in bacterial lipids at ambient temperature. With a novel membrane-anchored biradical and purpose-built hardware a 17-fold enhancement in NMR signal intensity is obtained by DNP which is one of the best obtained for a truly static matrix-free system. Furthermore, a membrane anchor sequence encompassing
    Mots-clés : angle, enhanced nmr, high-frequency, lipid-bilayers, magnetic-resonance, polarizing agents, polypeptides, protein, solids, Temperature.

  • Sauvee, C.; Casano, G.; Abel, S.; Rockenbauer, A.; Akhmetzyanov, D.; Karoui, H.; Siri, D.; Aussenac, F.; Maas, W.; Weber, R. T.; Prisner, T.; Rosay, M.; Tordo, P.; Ouari, O. Chemistry-a European Journal 2016, 22 (16), 5598-5606.
    Résumé : A series of 18 nitroxide biradicals derived from bTurea has been prepared, and their enhancement factors (H-1) in cross-effect dynamic nuclear polarization (CEDNP) NMR experiments at 9.4 and 14.1T and 100K in a DNP-optimized glycerol/water matrix (DNP juice) have been studied. We observe that (H-1) is strongly correlated with the substituents on the polarizing agents, and its trend is discussed in terms of different molecular parameters: solubility, average e-e distance, relative orientation of the nitroxide moieties, and electron spin relaxation times. We show that too short an e-e distance or too long a T-1e can dramatically limit (H-1). Our study also shows that the molecular structure of AMUPol is not optimal and its (H-1) could be further improved through stronger interaction with the glassy matrix and a better orientation of the TEMPO moieties. A new AMUPol derivative introduced here provides a better (H-1) than AMUPol itself (by a factor of ca. 1.2).
    Mots-clés : broadened esr line, dinitroxides, glassy solvents, high-frequency, magnetic-resonance, metal-organic frameworks, molecular dynamics, nitroxyl radicals, organic synthesis, polarizing agents, sensitivity-enhanced nmr, sn-beta zeolite, solid-state nmr, solvent effects, theoretical aspects.

  • Zielonka, J.; Zielonka, M.; VerPlank, L.; Cheng, G.; Hardy, M.; Ouari, O.; Ayhan, M. M.; Podsiadly, R.; Sikora, A.; Lambeth, J. D.; Kalyanaraman, B. Journal of Biological Chemistry 2016, 291 (13), 7029-7044.
    Résumé : Using high throughput screening-compatible assays for superoxide and hydrogen peroxide, we identified potential inhibitors of the NADPH oxidase (Nox2) isoform from a small library of bioactive compounds. By using multiple probes (hydroethidine, hydropropidine, Amplex Red, and coumarin boronate) with well defined redox chemistry that form highly diagnostic marker products upon reaction with superoxide (O2 arrange=stack.), hydrogen peroxide (H2O2), and peroxynitrite (ONOO-), the number of false positives was greatly decreased. Selected hits for Nox2 were further screened for their ability to inhibit ONOO- formation in activated macrophages. A new diagnostic marker product for ONOO- is reported. We conclude that the newly developed high throughput screening/reactive oxygen species assays could also be used to identify potential inhibitors of ONOO- formed from Nox2-derived O2. and nitric oxide synthase-derived nitric oxide.
    Mots-clés : biology, cancer, cells, disease, extracellular-superoxide, fluorescence probes, high throughput screening (HTS), in-vivo, NADPH oxidase inhibitors, NADPH oxidase, nitric-oxide, nox inhibitors, reactive nitrogen species (RNS), reactive oxygen, reactive oxygen species (ROS), superoxide ion, therapeutic targets.
2015

  • Ayhan, M. M.; Karoui, H.; Hardy, M.; Rockenbauer, A.; Charles, L.; Rosas, R.; Udachin, K.; Tordo, P.; Bardelang, D.; Ouari, O. Journal of the American Chemical Society 2015, 137 (32), 10238-10245.
    Résumé : We describe a photochemical method to introduce a single alcohol function directly on cucurbit[n]urils (n = 5, 6, 7, 8) with conversions of the order 95-100% using hydrogen peroxide and UV light. The reaction was easily scaled up to 1 g for CB[6] and CB[7]. Spin trapping of cucurbituril radicals combined with MS experiments allowed us to get insights about the reaction mechanism and characterize CB [5], CB[6], CB[7], and C13[8] monofunctional compounds. Experiments involving O-18 isotopically labeled water indicated that the mechanism was complex and showed signs of both radical and ionic intermediates. DFT calculations allowed estimating the Bond Dissociation Energies (BDEs) of each hydrogen atom type in the CB series, providing an explanation of the higher reactivity of the "equatorial" C-H position of CB[n] compounds. These results also showed that, for CB [8], direct functionalization on the cucurbituril skeleton is more difficult because one of the methylene hydrogen atoms (H-b) has its BDE lowering within the series and coming close to that of H-c, thus opening the way to other types of free radicals generated on the CB[8] skeleton leading to several side products. Yet CB[5]-(OH)(1) and CB[8]-(OH)(1), the first CB[8] derivative, were obtained in excellent yields thanks to the soft method presented here.

  • Ayhan, M. M.; Casano, G.; Karoui, H.; Rockenbauer, A.; Monnier, V.; Hardy, M.; Tordo, P.; Bardelang, D.; Ouari, O. Chemistry (Weinheim an der Bergstrasse, Germany) 2015, 21 (46).
    Résumé : Host-Guest Systems After extensive work toward the creation of molecular capsules in which two cavitands self-assemble to produce a constricted space amenable to guest encapsulation, a water-soluble version of Rebek's capsules was recently introduced. By combining EPR and NMR spectroscopy and ESI-MS, O. Ouari and co-workers have investigated the inner space metrics (width and height) of these supramolecular objects in water, as well as their binding constants and guest rotational dynamics. Nitroxides have been shown to be powerful probes affording a wealth of information. Read more in the Full Paper on page 16404 ff.

  • Ayhan, M. M.; Casano, G.; Karoui, H.; Rockenbauer, A.; Monnier, V.; Hardy, M.; Tordo, P.; Bardelang, D.; Ouari, O. Chemistry-a European Journal 2015, 21 (46), 16404-+.
    Résumé : Nitroxide free radicals have been used to study the inner space of one of Rebek's water-soluble capsules. EPR and (HNMR)-H-1 spectroscopy, ESI-MS, and DFT calculations showed a preference for the formation of 1:2 complexes. EPR titrations allowed us to determine binding constants (K-a) in the order of 10(7)M(-2). EPR spectral-shape analysis provided information on the guest rotational dynamics within the capsule. The interplay between optimum hydrogen bonding upon capsule formation and steric strain for guest accommodation highlights some degree of flexibility for guest inclusion, particularly at the center of the capsule where the hydrogen bond seam can be barely distorted or slightly disturbed.

  • Cheng, G.; Zielonka, J.; McAllister, D.; Hardy, M.; Ouari, O.; Joseph, J.; Dwinell, M. B.; Kalyanaraman, B. Cancer Letters 2015, 365 (1), 96-106.
    Résumé : One of the proposed mechanisms for tumor proliferation involves redox signaling mediated by reactive oxygen species such as superoxide and hydrogen peroxide generated at moderate levels. Thus, the antiproliferative and anti-tumor effects of certain antioxidants were attributed to their ability to mitigate intracellular reactive oxygen species (ROS). Recent reports support a role for mitochondrial ROS in stimulating tumor cell proliferation. In this study, we compared the antiproliferative effects and the effects on mitochondrial bioenergetic functions of a mitochondria-targeted cationic carboxyproxyl nitroxide (Mito-CP), exhibiting superoxide dismutase (SOD)-like activity and a synthetic cationic acetamide analog (Mito-CP-Ac) lacking the nitroxide moiety responsible for the SOD activity Results indicate that both Mito-CP and Mito-CP-Ac potently inhibited tumor cell proliferation. Both compounds altered mitochondrial and glycolytic functions, and intracellular citrate levels. Both Mito-CP and Mito-CP-Ac synergized with 2-deoxy-glucose (2-DG) to deplete intracellular ATP, inhibit cell proliferation and induce apoptosis in pancreatic cancer cells. We conclude that mitochondria-targeted cationic agents inhibit tumor proliferation via modification of mitochondrial bioenergetics pathways rather than by dismutating and detoxifying mitochondrial superoxide. (C) 2015 Elsevier Ireland Ltd. All rights reserved.

  • Debowska, K.; Debski, D.; Hardy, M.; Jakubowska, M.; Kalyanaraman, B.; Marcinek, A.; Michalski, R.; Michalowski, B.; Ouari, O.; Sikora, A.; Smulik, R.; Zielonka, J. Pharmacological Reports 2015, 67 (4), 756-764.
    Résumé : Over the last 40 years, there has been tremendous progress in understanding the biological reactions of reactive oxygen species (ROS) and reactive nitrogen species (RNS). It is widely accepted that the generation of ROS and RNS is involved in physiological and pathophysiological processes. To understand the role of ROS and RNS in a variety of pathologies, the specific detection of ROS and RNS is fundamental. Unfortunately, the intracellular detection and quantitation of ROS and RNS remains a challenge. In this short review, we have focused on the mechanistic and quantitative aspects of their detection with the use of selected fluorogenic probes. The challenges, limitations and perspectives of these methods are discussed. (C) 2015 Institute of Pharmacology, Polish Academy of Sciences. Published by Elsevier Sp. z o.o. All rights reserved.

  • Kaur, H.; Lakatos, A.; Spadaccini, R.; Vogel, R.; Hoffmann, C.; Becker-Baldus, J.; Ouari, O.; Tordo, P.; Mchaourab, H.; Glaubitz, C. Biological Chemistry 2015, 396 (9-10), 1135-1149.
    Résumé : ATP binding cassette (ABC) transporters form a superfamily of integral membrane proteins involved in translocation of substrates across the membrane driven by ATP hydrolysis. Despite available crystal structures and extensive biochemical data, many open questions regarding their transport mechanisms remain. Therefore, there is a need to explore spectroscopic techniques such as solid state NMR in order to bridge the gap between structural and mechanistic data. In this study, we investigate the feasibility of using Escherichia coli MsbA as a model ABC transporter for solid state NMR studies. We show that optimised solubilisation and reconstitution procedures enable preparing stable and homogenous protein samples. Depending on the duration of solubilisation, MsbA can be obtained in either an apo- or in a native lipid A bound form. Building onto these optimisations, the first promising MAS-NMR spectra with narrow lines have been recorded. However, further sensitivity improvements are required so that complex NMR experiments can be recorded within a reasonable amount of time. We therefore demonstrate the usability of paramagnetic doping for rapid data acquisition and explore dynamic nuclear polarisation as a method for general signal enhancement. Our results demonstrate that solid state NMR provides an opportunity to address important biological questions related to complex mechanisms of ABC transporters.

  • Le, D.; Ziarelli, F.; Phan, T. N. T.; Mollica, G.; Thureau, P.; Aussenac, F.; Ouari, O.; Gigmes, D.; Tordo, P.; Viel, S. Macromolecular Rapid Communications 2015, 36 (15), 1416-1421.
    Résumé : High-field dynamic nuclear polarization (DNP) has emerged as a powerful technique for improving the sensitivity of solid-state NMR (SSNMR), yielding significant sensitivity enhancements for a variety of samples, including polymers. Overall, depending upon the type of polymer, the molecular weight, and the DNP sample preparation method, sensitivity enhancements between 5 and 40 have been reported. These promising enhancements remain, however, far from the theoretical maximum (>1000). Crucial to the success of DNP SSNMR is the DNP signal enhancement (epsilon(DNP)), which is the ratio of the NMR signal intensities with and without DNP. It is shown here that, for polymers exhibiting high affinity toward molecular oxygen (e.g., polystyrene), removing part of the absorbed (paramagnetic) oxygen from the solid-state samples available as powders (instead of dissolved or dispersed in a solvent) increases proton nuclear relaxation times and epsilon(DNP), hereby providing up to a two-fold sensitivity increase (i.e., a four-fold reduction in experimental time).

  • Lelli, M.; Chaudhari, S. R.; Gajan, D.; Casano, G.; Rossini, A. J.; Ouari, O.; Tordo, P.; Lesage, A.; Emsley, L. Journal of the American Chemical Society 2015, 137 (46), 14558-14561.
    Résumé : Efficient dynamic nuclear Efficient dynamic nuclear polarization (DNP) in solids, which enables very high sensitivity NMR experiments, is currently limited to temperatures of around 100 K and below. Here we show how by choosing an adequate solvent, 1H cross effect DNP enhancements of over 80 can be obtained at 240 K. To achieve this we use the biradical TEKPol dissolved in a glassy phase of ortho-terphenyl (OTP). We study the solvent DNP enhancement of both TEKPol and BDPA in OTP in the range from 100 to 300 K at 9.4 and 18.8 T. Surprisingly, we find that the DNP enhancement decreases only relatively slowly for temperatures below the glass transition of OTP (T-g = 243 K), and 1H enhancements around 1520 at ambient temperature can be observed. We use this to monitor molecular dynamic transitions in the pharmaceutically relevant solids Ambroxol and Ibuprofen.

  • Maciejko, J.; Mehler, M.; Kaur, J.; Lieblein, T.; Morgner, N.; Ouari, O.; Tordo, P.; Becker-Baldus, J.; Glaubitz, C. Journal of the American Chemical Society 2015, 137 (28), 9032-9043.
    Résumé : Membrane proteins often form oligomeric complexes within the lipid bilayer, but factors controlling their assembly are hard to predict and experimentally difficult to determine. An understanding Of protein-protein interactions within the lipid bilayer is however required in order to elucidate the role of oligomerization for their functional mechanism and stabilization. Here, we demonstrate for the pentameric, heptahelical membrane protein green proteorhodopsin that solid-state NMR could identify specific interactions at the protomer interfaces, if the sensitivity is enhanced by dynamic nuclear polarization. For this purpose, differently labeled protomers have been assembled into the hill pentamer complex embedded within the lipid bilayer. We show for this proof of concept that One specific salt bridge determines the formation of pentamers or hexamers. Data are supported by laser-induced liquid bead ion desorption mass spectrometry and by blue native polyacrylamide gel electrophoresis analysis. The presented approach is universally applicable and opens the door toward analyzing membrane protein interactions within homo-oligomers directly in the membrane.

  • Salnikov, E. S.; Sarrouj, H.; Reiter, C.; Aisenbrey, C.; Purea, A.; Aussenac, F.; Ouari, O.; Tordo, P.; Fedotenko, I.; Engelke, F.; Bechinger, B. Journal of Physical Chemistry B 2015, 119 (46), 14574-14583.
    Résumé : Dynamic nuclear polarization has been developed to overconie the limitations of the inherently low signal intensity of NMR spectrostopy. This technique promises to be particularlynsefol for solid-state NMR spectroscopy where the. signals are broadened over d larger frequency range and most investigations rely on recording low gamma nuclei. To extend the range of-possible investigations, a triple-resonance flat-coil solid-state NMR probe is presented with microwave irradiation capacities allowing the investigation of Static samples at tempetatures of 100 K including supported lipid bilayets. The probe performance allows for two-dimensional separated local field experiments with high-power Lee-Goldberg decoupling and cross-polarization under simultaneous irradiation from,gyrotron micro-wave generator. Efficient cooling of the sample turned out to be essential for best enhancements and line shape and necessitated the development of a dedicated cooling chamber. Furthermore, a new membrane-anchoted biradical is presented, and the geometry of supported membranes was,optimized not only for good Membrane alignment, handling) stability, and filling factor of the coil but also for heat and microwave dissipation. Enhancement factors of 17-fold were obtained, and a two-dimensional PISEMA spectrum of a transmembrane helical peptide was obtained in lessthan 2 h.

  • van der Cruijsen, E. A. W.; Koers, E. J.; Sauvee, C.; Hulse, R. E.; Weingarth, M.; Ouari, O.; Perozo, E.; Tordo, P.; Baldus, M. Chemistry-a European Journal 2015, 21 (37), 12971-12977.
    Résumé : Dynamic nuclear polarization (DNP) has been shown to greatly enhance spectroscopic sensitivity, creating novel opportunities for NMR studies on complex and large molecular assemblies in life and material sciences. In such applications, however, site-specificity and spectroscopic resolution become critical factors that are usually difficult to control by current DNP-based approaches. We have examined in detail the effect of directly attaching mono- or biradicals to induce local paramagnetic relaxation effects and, at the same time, to produce sizable DNP enhancements. Using a membrane-embedded ion channel as an example, we varied the degree of paramagnetic labeling and the location of the DNP probes. Our results show that the creation of local spin clusters can generate sizable DNP enhancements while preserving the intrinsic benefits of paramagnetic relaxation enhancement (PRE)-based NMR approaches. DNP using chemical labeling may hence provide an attractive route to introduce molecular specificity into DNP studies in life science applications and beyond.

  • Ysacco, C.; Giorgi, M.; Tordo, P.; Bardelang, D.; Ouari, O. Canadian Journal of Chemistry 2015, 93 (8), 920-924.
    Résumé : A dinitroxide biradical (dCdO) has the peculiar tendency to crystallize in non-centrosymmetric polar space groups either as a stable dichloromethane solvate or as a pure phase. The molecules arrange with no compensation of their dipolar moment resulting in net macroscopic polar order.
2014

  • Abbas, K.; Hardy, M.; Poulhes, F.; Karoui, H.; Tordo, P.; Ouari, O.; Peyrot, F. Free Radical Biology and Medicine 2014, 71, 281-290.
    Résumé : Reactive oxygen species (ROS), including superoxide anion and hydrogen peroxide (H2O2), have a diverse array of physiological and pathological effects within living cells depending on the extent, timing, and location of their production. For measuring ROS production in cells, the ESR spin trapping technique using cyclic nitrones distinguishes itself from other methods by its specificity for superoxide and hydroxyl radical. However, several drawbacks, such as the low spin trapping rate and the spontaneous and cell-enhanced decomposition of the spin adducts to ESR-silent products, limit the application of this method to biological systems. Recently, new cyclic nitrones bearing a triphenylphosphonium (Mito-DIPPMPO) or a permethylated beta-cyclodextrin moiety (CD-DIPPMPO) have been synthesized and their spin adducts demonstrated increased stability in buffer. In this study, a comparison of the spin trapping efficiency of these new compounds with commonly used cyclic nitrone spin traps, i.e., 5,5-dimethyl-1-pyrroline N-oxide (DMPO), and analogs BMPO, DEPMPO, and DIPPMPO, was performed on RAW 264.7 macrophages stimulated with phorbol 12-myristate 13-acetate. Our results show that Mito-DIPPMPO and CD-DIPPMPO enable a higher detection of superoxide adduct, with a low (if any) amount of hydroxyl adduct. CD-DIPPMPO, especially, appears to be a superior spin trap for extracellular superoxide detection in living macrophages, allowing measurement of superoxide production in unstimulated cells for the first time. The main rationale put forward for this extreme sensitivity is that the extracellular localization of the spin trap prevents the reduction of the spin adducts by ascorbic acid and glutathione within cells. (C) 2014 Elsevier Inc. All rights reserved.

  • Bardelang, D.; Casano, G.; Poulhes, F.; Karoui, H.; Filippini, J.; Rockenbauer, A.; Rosas, R.; Monnier, V.; Siri, D.; Gaudel-Siri, A.; Ouari, O.; Tordo, P. Journal of the American Chemical Society 2014, 136 (50), 17570-17577.
    Résumé : The flexible tetranitroxide 4T has been prepared and was shown to exhibit a nine line EPR spectrum in water, characteristic of significant through space spin exchange (J(ij)) between four electron spins interacting with four nitrogen nuclei (J(ij) >= a(N)). Addition of CB[8] to 4T decreases dramatically all the J(ij) couplings, and the nine line spectrum is replaced by the characteristic three line spectrum of a mononitroxide. The supramolecular association between 4T and CB[8] involves a highly cooperative asymmetric complexation by two CB[8] (K-1 = 4027 M-1; K-2 = 202 800 M-1; alpha = 201) leading to a rigid complex with remote nitroxide moieties. The remarkable enhancement for the affinity of the second CB[8] corresponds to an allosteric interaction energy of approximate to 13 kJ mol(-1), which is comparable to that of the binding of oxygen by hemoglobin. These results are confirmed by competition and reduction experiments, DFT and molecular dynamics calculations, mass spectrometry, and liquid state NMR of the corresponding reduced complex bearing hydroxylamine moieties. This study shows that suitably designed molecules can generate allosteric complexation with CB[8]. The molecule must (i) carry several recognizable groups for CB[8] and (ii) be folded so that the first binding event reorganizes the molecule (unfold) for a better subsequent recognition. The presence of accessible protonable amines and H-bond donors to fit with the second point are also further stabilizing groups of CB[8] complexation. In these conditions, the spin exchange coupling between four radicals has been efficiently and finely tuned and the resulting allosteric complexation induced a dramatic stabilization enhancement of the included paramagnetic moieties in highly reducing conditions through the formation of the supramolecular 4T@CB[8](2) complex.

  • Bardelang, D.; Giorgi, M.; Hornebecq, V.; Stepanov, A.; Hardy, M.; Rizzato, E.; Monnier, V.; Zaman, M. B.; Chan, G.; Udachin, K.; Enright, G.; Tordo, P.; Ouari, O. Crystal Growth & Design 2014, 14 (2), 467-476.
    Résumé : The dinitroxide bis(TEMPO) bisketal (bTbk) was shown to crystallize into open frameworks whose structures were determined by single-crystal X-ray diffraction. We show that bTbk can be used as a supramolecular building block for the hosting of a plethora of guests inside the ID channels of its paramagnetic framework, including other radicals such as TEMPO or 2-azaadamantane-N-oxyl. C-60 and C-70 were also found to be easily included in this open framework during its crystallization. This resulted in well-defined, nanostructured assemblies of composite radical crystals (bTbk/toluene/C-60 or C-70) or (bTbk/toluene/TEMPO) by a very simple dissolution/crystallization process with tunable guest content. Selective C-60 extraction was also demonstrated directly from fullerene soot.

  • Beziere, N.; Hardy, M.; Poulhes, F.; Karoui, H.; Tordo, P.; Ouari, O.; Frapart, Y. - M.; Rockenbauer, A.; Boucher, J. - L.; Mansuy, D.; Peyrot, F. Free Radical Biology and Medicine 2014, 67, 150-158.
    Résumé : Reactive oxygen species are by-products of aerobic metabolism involved in the onset and evolution of various pathological conditions. Among them, the superoxide radical is of special interest as the origin of several damaging species such as H2O2, hydroxyl radical, or peroxynitrite (ONOO-). Spin trapping coupled with ESR is a method of choice to-characterize these species in chemical and biological systems and the metabolic stability of the spin adducts derived from reaction of superoxide and hydroxyl radicals with nitrones is the main limit to the in vivo application of the method. Recently, new cyclic nitrones bearing a triphenylphosphonium or permethylated beta-cyclodextrin moiety have been synthesized and their spin adducts demonstrated increased stability in buffer. In this article, we studied the stability of the superoxide adducts of four new cyclic nitrones in the presence of liver subcellular fractions and biologically relevant reductants using an original setup combining a stopped-flow device and an ESR spectrometer. The kinetics of disappearance of the spin adducts were analyzed using an appropriate simulation program. Our results highlight the interest of the new spin trapping agents CD-DEPMPO and CD-DIPPMPO for specific detection of superoxide with high stability of the superoxide adducts in the presence of liver microsomes. (C) 2013 Elsevier Inc. All rights reserved.

  • Conley, M. P.; Rossini, A. J.; Comas-Vives, A.; Valla, M.; Casano, G.; Ouari, O.; Tordo, P.; Lesage, A.; Emsley, L.; Coperet, C. Physical Chemistry Chemical Physics 2014, 16 (33), 17822-17827.
    Résumé : Grafting reactive molecular complexes on dehydroxylated amorphous silica is a strategy to develop "single-site" heterogeneous catalysts. In general, only the reactivity of isolated silanols is invoked for silica dehydroxylated at 700 degrees C ([SiO2-700]), though ca. 10% of the surface silanols are in fact geminal Q(2)-silanols. Here we report the reaction of allyltributylstannane with (SiO2-700] and find that the geminal Q(2)-silanols react to form products that would formally arise from vicinal Q(3)-silanols that are not present on (SiO2-700], indicating that a surface rearrangement occurs. The reorganization of the silica surface is unique to silica dehydroxylated at 700 degrees C or above. The findings were identified using Dynamic Nuclear Polarization Surface Enhanced NMR Spectroscopy (DNP SENS) combined with DFT calculations.

  • Hardy, M.; Poulhes, F.; Rizzato, E.; Rockenbauer, A.; Banaszak, K.; Karoui, H.; Lopez, M.; Zielonka, J.; Vasquez-Vivar, J.; Sethumadhavan, S.; Kalyanaraman, B.; Tordo, P.; Ouari, O. Chemical Research in Toxicology 2014, 27 (7), 1155-1165.
    Résumé : Development of reliable methods and site-specific detection of free radicals is an active area of research. Here, we describe the synthesis and radical-trapping properties of new derivatives of DEPMPO and DIPPMPO, bearing a mitochondria-targeting triphenylphosphonium cationic moiety or guanidinium cationic group. All of the spin traps prepared have been observed to efficiently trap superoxide radical anions in a cell-free system. The superoxide spin adducts exhibited similar spectral properties, indicating no significant differences in the geometry of the cyclic nitroxide moieties of the spin adducts. The superoxide adduct stability was measured and observed to be highest (t(1/2) = 73 min) for DIPPMPO nitrone linked to triphenylphosphonium moiety via a short carbon chain (Mito-DIPPMPO). The experimental results and DFT quantum chemical calculations indicate that the cationic property of the triphenylphosphonium group may be responsible for increased superoxide trapping efficiency and adduct stability of Mito-DIPPMPO, as compared to the DIPPMPO spin trap. The studies of uptake of the synthesized traps into isolated mitochondria indicated the importance of both cationic and lipophilic properties, with the DEPMPO nitrone linked to the triphenylphosphonium moiety via a long carbon chain (Mito(10)-DEPMPO) exhibiting the highest mitochondrial uptake. We condude that, of the synthesized traps, Mito-DIPPMPO and Mito(10)-DEPMPO are the best candidates for potential mitochondria-specific spin traps for use in biologically relevant systems.

  • Karoui, H.; Nsanzumuhire, C.; Le Moigne, F.; Hardy, M.; Siri, D.; Derat, E.; Rockenbauer, A.; Ouari, O.; Tordo, P. Chemistry-a European Journal 2014, 20 (14), 4064-4071.
    Résumé : The 5-diethoxyphosphonyl-5-methyl-1-pyrroline N-oxide superoxide spin adduct (DEPMPOOOH) is much more persistent (about 15 times) than the 5,5-dimethyl-1-pyrroline N-oxide superoxide spin adduct (DMPOOOH). The diethoxyphosphonyl group is bulkier than the methyl group and its electron-withdrawing effect is much stronger. These two factors could play a role in explaining the different half-lifetimes of DMPOOOH and DEPMPOOOH. The trifluoromethyl and the diethoxyphosphonyl groups show similar electron-withdrawing effects but have different sizes. We have thus synthesized and studied 5-methyl-5-trifluoromethyl-1-pyrroline N-oxide (5-TFDMPO), a new trifluoromethyl analogue of DMPO, to compare its spin-trapping performance with those of DMPO and DEPMPO. 5-TFDMPO was prepared in a five-step sequence by means of the Zn/AcOH reductive cyclization of 5,5,5-trifluoro-4-methyl-4-nitropentanal, and the geometry of the molecule was estimated by using DFT calculations. The spin-trapping properties were investigated both in toluene and in aqueous buffer solutions for oxygen-, sulfur-, and carbon-centered radicals. All the spin adducts exhibit slightly different fluorine hyperfine coupling constants, thereby suggesting a hindered rotation of the trifluoromethyl group, which was confirmed by variable-temperature EPR studies and DFT calculations. In phosphate buffer at pH7.4, the half-life of 5-TFDMPOOOH is about three times shorter than for DEPMPOOOH and five times longer than for DMPOOOH. Our results suggest that the stabilization of the superoxide adducts comes from a delicate balance between steric, electronic, and hydrogen-bonding effects that involve the group, the hydroperoxyl moiety, and the nitroxide.

  • Koers, E. J.; van der Cruijsen, E. A. W.; Rosay, M.; Weingarth, M.; Prokofyev, A.; Sauvee, C.; Ouari, O.; van der Zwan, J.; Pongs, O.; Tordo, P.; Maas, W. E.; Baldus, M. Journal of Biomolecular Nmr 2014, 60 (2-3), 157-168.
    Résumé : Dynamic nuclear polarization (DNP) has become a powerful method to enhance spectroscopic sensitivity in the context of magnetic resonance imaging and nuclear magnetic resonance spectroscopy. We show that, compared to DNP at lower field (400 MHz/263 GHz), high field DNP (800 MHz/527 GHz) can significantly enhance spectral resolution and allows exploitation of the paramagnetic relaxation properties of DNP polarizing agents as direct structural probes under magic angle spinning conditions. Applied to a membrane-embedded K+ channel, this approach allowed us to refine the membrane-embedded channel structure and revealed conformational substates that are present during two different stages of the channel gating cycle. High-field DNP thus offers atomic insight into the role of molecular plasticity during the course of biomolecular function in a complex cellular environment.

  • Kubicki, D. J.; Rossini, A. J.; Purea, A.; Zagdoun, A.; Ouari, O.; Tordo, P.; Engelke, F.; Lesage, A.; Emsley, L. Journal of the American Chemical Society 2014, 136 (44), 15711-15718.
    Résumé : There is currently great interest in understanding the limits on NMR signal enhancements provided by dynamic nuclear polarization (DNP), and in particular if the theoretical maximum enhancements can be achieved. We show that over a 2-fold improvement in cross-effect DNP enhancements can be achieved in MAS experiments on frozen solutions by simply incorporating solid particles into the sample. At 9.4 T and similar to 105 K, enhancements up to epsilon(H) = 515 are obtained in this way, corresponding to 78% of the theoretical maximum. We also underline that degassing of the sample is important to achieve highest enhancements. We link the amplification effect to the dielectric properties of the solid material, which probably gives rise to scattering, diffraction, and amplification of the microwave field in the sample. This is substantiated by simulations of microwave propagation. A reduction in sample heating at a given microwave power also likely occurs due to reduced dielectric loss. Simulations indicate that the microwave field (and thus the DNP enhancement) is inhomogeneous in the sample, and we deduce that in these experiments between 5 and 10% of the solution actually yields the theoretical maximum signal enhancement of 658. The effect is demonstrated for a variety of particles added to both aqueous and organic biradical solutions.


  • Le, D.; Casano, G.; Phan, T. N. T.; Ziarelli, F.; Ouari, O.; Aussenac, F.; Thureau, P.; Mollica, G.; Gigmes, D.; Tordo, P.; Viel, S. Macromolecules 2014, 47 (12), 3909-3916.
    Résumé : This work compares the overall sensitivity enhancements provided by dynamic nuclear polarization (DNP) for the solid-state NMR characterization of polymer samples doped with biradicals and prepared either by film casting (FC), or by glass forming (GF) using 1,1,2,2-tetrachloroethane as the solvent. Analysis of amorphous and semicrystalline polymers (polystyrene, poly(ethylene oxide), polylactide, poly(methyl methacrylate)) of varying molecular weights showed that GF provided larger sensitivity enhancements than FC but yielded DNP-enhanced 13C CPMAS spectra of lower resolution for semicrystalline polymers, owing to line-broadening due to conformational distribution of the polymer chains in frozen solution. Moreover, use of deuterated solvents significantly reduced the intensity of the solvent signals in the DNP-enhanced 13C CPMAS spectra of polymers prepared by GF, while preserving the sensitivity enhancement observed for the polymer signals. For the polymers investigated here, both FC and GF performed better than incipient wetness impregnation, yielding overall sensitivity enhancements between 5 and 40.

  • Lelli, M.; Rossini, A. J.; Casano, G.; Ouari, O.; Tordo, P.; Lesage, A.; Emsley, L. Chemical Communications 2014, 50 (71), 10198-10201.
    Résumé : We show how large DNP enhancements of NMR signals can be obtained from several hydrophobic radicals that are solubilised in aqueous environments by a variety of biologically compatible neutral amphiphiles. In particular we show that the bi-radical TEKPOL can be incorporated into micelles formed by the surfactant polysorbate 80 (Tween-80), where we obtain large DNP enhancements (similar to 60) at 9.4 T and similar to 100 K.


  • Mollica, G.; Le, D.; Ziarelli, F.; Casano, G.; Ouari, O.; Phan, T. N. T.; Aussenac, F.; Thureau, P.; Gigmes, D.; Tordo, P.; Viel, S. ACS Macro Letters 2014, 3 (9), 922-925.
    Résumé : High-field dynamic nuclear polarization (DNP) may enhance the sensitivity of solid-state NMR experiments on a wide range of systems, including synthetic polymers, owing to the transfer of electron spin polarization from radicals to nuclei upon microwave irradiation (usually at cryogenic temperatures). Provided that the radicals are homogeneously dispersed in the sample, a uniform DNP enhancement is expected for all the signals in the 13C cross-polarization magic angle spinning (CPMAS) spectrum. Here, we show that, in the case of methyl group containing polymers, a change in the cross-polarization (CP) dynamics induced by the moderate increase in sample temperature due to microwave irradiation may lead to the observation of apparent nonuniform enhancements in the DNP-enhanced 13C CPMAS spectra. This peculiar behavior should be accounted for when measuring 13C CP DNP enhancements on polymer materials, especially for heterogeneous polymer samples (for which truly nonuniform DNP enhancements could potentially be detected), or when quantitative results are sought.
2013

  • Bardelang, D.; Giorgi, M.; Pardanaud, C.; Hornebecq, V.; Rizzato, E.; Tordo, P.; Ouari, O. Chemical Communications 2013, 49 (34), 3519-3521.
    Résumé : The dinitroxide biradical bTbk crystallizes forming hexagonal open frameworks with one-dimensional corrugated channels filled with crystallization solvent. The large pockets constitutive of the channels allowed the inclusion of C-60 in the paramagnetic network. The rapidity and high fidelity of crystal growth were used to prepare isostructural multilayer host-guest crystals successively stained with C-60.

  • Cheng, T.; Mishkovsky, M.; Bastiaansen, J. A. M.; Ouari, O.; Hautle, P.; Tordo, P.; van den Brandt, B.; Comment, A. Nmr in Biomedicine 2013, 26 (11), 1582-1588.
    Résumé : Hyperpolarized magnetic resonance via dissolution dynamic nuclear polarization necessitates the transfer of the hyperpolarized molecules from the polarizer to the imager prior to in vivo measurements. This process leads to unavoidable losses in nuclear polarization, which are difficult to evaluate once the solution has been injected into an animal. We propose a method to measure the polarization of the hyperpolarized molecules inside the imager bore, 3 s following dissolution, at the time of the injection, using a precise quantification of the infusate concentration. This in situ quantification allows for distinguishing between signal modulations related to variations in the nuclear polarization at the time of the injection and signal modulations related to physiological processes such as tissue perfusion. In addition, our method includes a radical scavenging process that leads to a minor reduction in sample concentration and takes place within a couple of seconds following the dissolution in order to minimize the losses due to the presence of paramagnetic polarizing agent in the infusate. We showed that proton exchange between vitamin C, the scavenging molecule and the deuterated solvent shortens the long carboxyl (13)C longitudinal relaxation time in [1-C-13]acetate. This additional source of dipolar relaxation can be avoided by using deuterated ascorbate. Overall, the method allows for a substantial gain in polarization and also leads to an extension of the time window available for in vivo measurements.

  • Conley, M. P.; Drost, R. M.; Baffert, M.; Gajan, D.; Elsevier, C.; Franks, W. T.; Oschkinat, H.; Veyre, L.; Zagdoun, A.; Rossini, A.; Lelli, M.; Lesage, A.; Casano, G.; Ouari, O.; Tordo, P.; Emsley, L.; Coperet, C.; Thieuleux, C. Chemistry-a European Journal 2013, 19 (37), 12234-12238.

  • Gajan, D.; Schwarzwaelder, M.; Conley, M. P.; Gruening, W. R.; Rossini, A. J.; Zagdoun, A.; Lelli, M.; Yulikov, M.; Jeschke, G.; Sauvee, C.; Ouari, O.; Tordo, P.; Veyre, L.; Lesage, A.; Thieuleux, C.; Emsley, L.; Coperet, C. Journal of the American Chemical Society 2013, 135 (41), 15459-15466.
    Résumé : Mesoporous hybrid silica-organic materials containing homogeneously distributed stable mono- or dinitroxide radicals covalently bound to the silica surface were developed as polarization matrixes for solid-state dynamic nuclear polarization (DNP) NMR experiments. For TEMPO-containing materials impregnated with water or 1,1,2,2-tetrachloroethane, enhancement factors of up to 36 were obtained at similar to 100 K and 9.4 T without the need for a glass-forming additive. We show that the homogeneous radical distribution and the subtle balance between the concentration of radical in the material and the fraction of radicals at a sufficient inter-radical distance to promote the cross-effect are the main determinants for the DNP enhancements we obtain. The material, as well as an analogue containing the poorly soluble biradical bTUrea, is used as a polarizing matrix for DNP NMR experiments of solutions containing alanine and pyruvic acid. The analyte is separated from the polarization matrix by simple filtration.

  • Mao, J.; Akhmetzyanov, D.; Ouari, O.; Denysenkov, V.; Corzilius, B.; Plackmeyer, J.; Tordo, P.; Prisner, T. F.; Glaubitz, C. Journal of the American Chemical Society 2013, 135 (51), 19275-19281.
    Résumé : Dynamic nuclear polarization (DNP) enhances the sensitivity of solid-state NMR (SSNMR) spectroscopy by orders of magnitude and, therefore, opens possibilities for novel applications from biology to materials science. This multitude of opportunities implicates a need for high-performance polarizing agents, which integrate specific physical and chemical features tailored for various applications. Here, we demonstrate that for the biradical bTbK in complex with captisol (CAP), a beta-cyclodextrin derivative, host-guest assembling offers a new and easily accessible approach for the development of new polarizing agents. In contrast to bTbK, the CAP-bTbK complex is water-soluble and shows significantly improved DNP performance compared to the commonly used DNP agent TOTAPOL. Furthermore, NMR and EPR data reveal improved electron and nuclear spin relaxation properties for bTbK within the host molecule. The numerous possibilities to functionalize host molecules will permit designing novel radical complexes targeting diverse applications.


  • Ouari, O.; Phan, T.; Ziarelli, F.; Casano, G.; Aussenac, F.; Thureau, P.; Gigmes, D.; Tordo, P.; Viel, S. ACS Macro Letters 2013, 2 (8), 715-719.
    Résumé : Dynamic nuclear polarization (DNP) is shown to greatly improve the solid-state nuclear magnetic resonance (SSNMR) analysis of synthetic polymers by allowing structural assignment of intrinsically diluted NMR signals, which are typically not detected in conventional SSNMR. Specifically, SSNMR and DNP SSNMR were comparatively used to study functional polymers for which precise structural elucidation of chain ends is essential to control their reactivity and to eventually obtain advanced polymeric materials of complex architecture. Results show that the polymer chain-end signals, while hardly observable in conventional SSNMR, could be clearly identified in the DNP SSNMR spectrum owing to the increase in sensitivity afforded by the DNP setup (a factor ?10 was achieved here), hence providing access to detailed structural characterization within realistic experimental times. This sizable gain in sensitivity opens new avenues for the characterization of ?smart? functional polymeric materials and new analytical perspectives in polymer science.

  • Sauvee, C.; Rosay, M.; Casano, G.; Aussenac, F.; Weber, R. T.; Ouari, O.; Tordo, P. Angewandte Chemie-International Edition 2013, 52 (41), 10858-10861.

  • Zagdoun, A.; Casano, G.; Ouari, O.; Schwaerzwalder, M.; Rossini, A. J.; Aussenac, F.; Yulikov, M.; Jeschke, G.; Coperet, C.; Lesage, A.; Tordo, P.; Emsley, L. Journal of the American Chemical Society 2013, 135 (34), 12790-12797.
    Résumé : A series of seven functionalized nitroxide biradicals (the bTbK biradical and six derivatives) are investigated as exogenous polarization sources for dynamic nuclear polarization (DNP) solid-state NMR at 9.4 T and with ca. 100 K sample temperatures. The impact of electron relaxation times on the DNP enhancement (epsilon) is examined, and we observe that longer inversion recovery and phase memory relaxation times provide larger E. All radicals are tested in both bulk 1,1,2,2-tetrachloroethane solutions and in mesoporous materials, and the difference in E between the two cases is discussed. The impact of the sample temperature and magic angle spinning frequency on epsilon is investigated for several radicals each characterized by a range of electron relaxation times. In particular, TEKPol, a bulky derivative of bTbK with a molecular weight of 905 g.mol(-1), is presented. Its high-saturation factor makes it a very efficient polarizing agent for DNP, yielding unprecedented proton enhancements of over 200 in both bulk and materials samples at 9.4 T and 100 K. TEKPol also yields encouraging enhancements of 33 at 180 K and 12 at 200 K, suggesting that with the continued improvement of radicals large e may be obtained at higher temperatures.
2012

  • Bardelang, D.; Giorgi, M.; Hornebecq, V.; Stepanov, A.; Rizzato, E.; Zaman, M. B.; Chan, G.; Ouari, O.; Tordo, P. Rsc Advances 2012, 2 (13), 5605-5609.
    Résumé : Toluene and 1-chloronaphthalene immobilization was observed when perturbing stimuli such as agitation or ultrasound are applied during the cooling of a hot supersaturated solution of the rigid dinitroxide bTbk. X-Ray diffraction and cryo-SEM on gels and solid samples (crystal, powder) confirmed the pivotal role of clathrate type microcrystals in the solvent immobilization process.

  • Dane, E. L.; Corzilius, B.; Rizzato, E.; Stocker, P.; Maly, T.; Smith, A. A.; Griffin, R. G.; Ouari, O.; Tordo, P.; Swagert, T. M. Journal of Organic Chemistry 2012, 77 (4), 1789-1797.
    Résumé : The synthesis and characterization of oxidized bis-thioketal-trispiro dinitroxide biradicals that orient the nitroxides in a rigid, approximately orthogonal geometry are reported. The biradicals show better performance as polarizing agents in dynamic nuclear polarization (DNP) NMR experiments as compared to biradicals lacking the constrained geometry. In addition, the biradicals display improved solubility in aqueous media due to the presence of polar sulfoxides. The results suggest that the orientation of the radicals is not dramatically affected by the oxidation state of the sulfur atoms in the biradical, and we conclude that a biradical polarizing agent containing a mixture of oxidation states can be used for improved solubility without a loss in performance.

  • Rossini, A. J.; Zagdoun, A.; Lelli, M.; Canivet, J.; Aguado, S.; Ouari, O.; Tordo, P.; Rosay, M.; Maas, W. E.; Coperet, C.; Farrusseng, D.; Emsley, L.; Lesage, A. Angewandte Chemie-International Edition 2012, 51 (1), 123-127.

  • Salnikov, E. S.; Ouari, O.; Koers, E.; Sarrouj, H.; Franks, T.; Rosay, M.; Pawsey, S.; Reiter, C.; Bandara, P.; Oschkinat, H.; Tordo, P.; Engelke, F.; Bechinger, B. Applied Magnetic Resonance 2012, 43 (1-2), 91-106.
    Résumé : Dynamic nuclear polarization (DNP)/solid-state nuclear magnetic resonance (NMR) spectroscopy bears great potential for the investigation of membrane-associated polypeptides which can often be produced only in small amounts and which need to be 'diluted' in lipid bilayer environments to adopt or maintain their functional structure. Here we present investigations using biradicals, such as TOTAPOL and bTbK, for solid-state NMR signal enhancement using DNP in the context of lipid membranes. By transferring polarization from electron to nuclear spins using microwave irradiation signal enhancement factors of up to 13 are obtained with TOTAPOL and up to 17 with bTbK. The possible reasons why these factors are below those obtained in glassy samples of bulk solvents (40-60 under similar conditions) are evaluated and discussed. In order to further ameliorate the enhancement factors the physico-chemical characteristics of TEMPOL, TOTAPOL, bTbK, and bCTbK, such as their partitioning between hydrophilic and hydrophobic solvents or their stability under different environmental conditions are presented. Finally, having provided proof-of-concept that DNP/solid-state NMR measurements can be performed with oriented membrane samples work in progress is presented on the development of a flat-coil probe for DNP/solid-state NMR experiments on oriented membranes.

  • Ysacco, C.; Karoui, H.; Casano, G.; Le Moigne, F.; Combes, S.; Rockenbauer, A.; Rosay, M.; Maas, W.; Ouari, O.; Tordo, P. Applied Magnetic Resonance 2012, 43 (1-2), 251-261.
    Résumé : We have prepared a series of dinitroxides and we investigated their properties as polarizing agents for solid-state nuclear magnetic resonance/dynamic nuclear polarization applications at 100 K, 9.34 T (263 GHz electron paramagnetic resonance and 400 MHz H-1 nuclear magnetic resonance). Our results show that a rigid structure with an orthogonal relative orientation of electron g tensors and the appropriate orientation of the two bonds are required to obtain maximum polarization enhancements. In addition, with dinitroxides exhibiting a long T (1e), the saturation of the irradiated electron spin packet is favored leading to more efficient dynamic nuclear polarization.

  • Zagdoun, A.; Casano, G.; Ouari, O.; Lapadula, G.; Rossini, A. J.; Lelli, M.; Baffert, M.; Gajan, D.; Veyre, L.; Maas, W. E.; Rosay, M.; Weber, R. T.; Thieuleux, C.; Coperet, C.; Lesage, A.; Tordo, P.; Emsley, L. Journal of the American Chemical Society 2012, 134 (4), 2284-2291.
    Résumé : A new nitroxide-based biradical having a long electron spin-lattice relaxation time (T-1e) has been developed as an exogenous polarization source for DNP solid-state NMR experiments. The performance of this new biradical is demonstrated on hybrid silica-based mesostructured materials impregnated with 1,1,2,2-tetrachloroethane radical containing solutions, as well as in frozen bulk solutions, yielding DNP enhancement factors (epsilon) of over 100 at a magnetic field of 9.4 T and sample temperatures of similar to 100 K. The effects of radical concentration on the DNP enhancement factors and on the overall sensitivity enhancements (Sigma(dagger)) are reported. The relatively high DNP efficiency of the biradical is attributed to an increased T-1e, which enables more effective saturation of the electron resonance. This new biradical is shown to outperform the polarizing agents used so far in DNP surface-enhanced NMR spectroscopy of materials, yielding a 113-fold increase in overall sensitivity for silicon-29 CPMAS spectra as compared to conventional NMR experiments at room temperature. This results in a reduction in experimental times by a factor >12 700, making the acquisition of C-13 and N-15 one- and two-dimensional NMR spectra at natural isotopic abundance rapid (hours). It has been used here to monitor a series of chemical reactions carried out on the surface functionalities of a hybrid organic-silica material.

  • Zagdoun, A.; Rossini, A. J.; Gajan, D.; Bourdolle, A.; Ouari, O.; Rosay, M.; Maas, W. E.; Tordo, P.; Lelli, M.; Emsley, L.; Lesage, A.; Coperet, C. Chemical Communications 2012, 48 (5), 654-656.
    Résumé : A series of non-aqueous solvents combined with the exogenous biradical bTbK are developed for DNP NMR that yield enhancements comparable to the best available water based systems. 1,1,2,2-tetrachloroethane appears to be one of the most promising organic solvents for DNP solid-state NMR. Here this results in a reduction in experimental times by a factor of 1000. These new solvents are demonstrated with the first DNP surface enhanced NMR characterization of an organometallic complex supported on a hydrophobic surface.

  • Zielonka, J.; Zielonka, M.; Sikora, A.; Adamus, J.; Joseph, J.; Hardy, M.; Ouari, O.; Dranka, B. P.; Kalyanaraman, B. Journal of Biological Chemistry 2012, 287 (5), 2984-2995.
    Résumé : Herein we describe a high-throughput fluorescence and HPLC-based methodology for global profiling of reactive oxygen and nitrogen species (ROS/RNS) in biological systems. The combined use of HPLC and fluorescence detection is key to successful implementation and validation of this methodology. Included here are methods to specifically detect and quantitate the products formed from interaction between the ROS/RNS species and the fluorogenic probes, as follows: superoxide using hydroethidine, peroxynitrite using boronate-based probes, nitric oxide-derived nitrosating species with 4,5-diaminofluorescein, and hydrogen peroxide and other oxidants using 10-acetyl-3,7-dihydroxyphenoxazine (Amplex (R) Red) with and without horseradish peroxidase, respectively. In this study, we demonstrate real-time monitoring of ROS/RNS in activated macrophages using high-throughput fluorescence and HPLC methods. This global profiling approach, simultaneous detection of multiple ROS/RNS products of fluorescent probes, developed in this study will be useful in unraveling the complex role of ROS/RNS in redox regulation, cell signaling, and cellular oxidative processes and in high-throughput screening of anti-inflammatory antioxidants.
2010

  • Gafurov, M.; Lyubenova, S.; Denysenkov, V.; Ouari, O.; Karoui, H.; Le Moigne, F.; Tordo, P.; Prisner, T. Applied Magnetic Resonance 2010, 37 (1-4), 505-514.
    Résumé : We have characterized the rigid binitroxide radical bis-TEMPO-bis-Ketal (bTbK) by continuous-wave (CW) and pulsed electron paramagnetic resonance (EPR) spectroscopy performed at X-band (9 GHz) and G-band (180 GHz) frequencies. bTbK has been successfully used for dynamic nuclear polarization (DNP)-enhanced solid-state nuclear magnetic resonance (SS-DNP) experiments based on the cross-effect, which involves two electrons and one nuclear spin, and gave very high signal enhancements. For a quantitative description of the polarization enhancements and their excitation frequency profile, a detailed information about the values and relative orientation of the magnetic hyperfine-, dipolar-, g-tensors and the exchange interaction of the two unpaired electron spins within the molecule is mandatory. We have determined these tensors and their relative orientation by CW-EPR spectra and pulsed electron double resonance experiments in frozen solution. The potential of using the cross-effect also for DNP in liquid solutions has been experimentally investigated by room-temperature high-field DNP experiments performed at 9.2 T.

  • Mieville, P.; Ahuja, P.; Sarkar, R.; Jannin, S.; Vasos, P. R.; Gerber-Lemaire, S.; Mishkovsky, M.; Comment, A.; Gruetter, R.; Ouari, O.; Tordo, P.; Bodenhausen, G. Angewandte Chemie-International Edition 2010, 49 (35), 6182-6185.

  • Salnikov, E.; Rosay, M.; Pawsey, S.; Ouari, O.; Tordo, P.; Bechinger, B. Journal of the American Chemical Society 2010, 132 (17), 5940-+.
    Résumé : Oriented membrane samples encompassing the biradical bTbK and a transmembrane peptide carrying a single (15)N labeled residue have been prepared on polymer sheets with sample geometries that fit into a 3.2 mm MAS rotor. The proton-decoupled (15)N cross-polarization spectra of the peptide were characterized by a single line at fast magic angle spinning speeds of similar to 8 kHz. Irradiating these samples with p-waves resulted in Dynamic Nuclear Polarization and a concomitant 18-fold signal enhancement which considerably shortened the NMR acquisition times. Furthermore, the sideband patterns of magic angle oriented sample spinning (MAOSS) solid-state NMR spectra at slow spinning speeds (similar to 1 kHz) are indicative that the lipids and peptides form well-oriented bilayers at 100 K despite the narrow inner diameter of the rotor (2.2 mm) and the presence of considerable amounts of biradicals. The DNP signal enhancement opens up enhanced possibilities for multidimensional solid-state NMR investigation of oriented membrane polypeptides.

  • Ysacco, C.; Rizzato, E.; Virolleaud, M. - A.; Karoui, H.; Rockenbauer, A.; Le Moigne, F.; Siri, D.; Ouari, O.; Griffin, R. G.; Tordo, P. Physical Chemistry Chemical Physics 2010, 12 (22), 5841-5845.
    Résumé : We have investigated the properties of a series of dinitroxides as polarizing agents for SS NMR/DNP applications. Our results clearly establish that an orthogonal relative orientation of electron g tensors is a crucial requirement to obtain high enhancement DNP factors. Moreover, the ratio of the proton Larmor frequency over the e(-)-e(-) dipolar coupling (omega(H)/omega(D)) influences the efficiency of the cross effect (CE) mechanism, thus the < Ree > distance between the unpaired electrons must be adapted to omega(H).