Nitrosonium Research Articles

Nitrosation of amides involves a pseudopericyclic 1,3-sigmatropic rearrangement.

Two possible pathways for the nitrosation of formamide and N-methyl formamide by nitrosonium ion (NO(+)) have been investigated at the B3LYP/6-31G(d,p) level. The key steps are pseudopericyclic 1,3-sigmatropic rearrangements to give the observed N-nitrosamides. The transition structures (8a and 8b) are close to planar on the amide moiety and have remarkably low barriers of only 6.6 and 4.8 kcal/mol from the lowest energy conformations of 6a and 6b, respectively. [reaction: see text]

A high-spin and durable polyradical: poly(4-diphenylaminium-1,2-phenylenevinylene).

A purely organic, high-spin, and durable polyradical molecule was synthesized: It is based on the non-Kekulé- and non-disjoint design of a pi-conjugated poly(1,2-phenylenevinylene) backbone pendantly 4-substituted with multiple robust arylaminium radicals. 4-N,N-Bis(4-methoxy- and -tert-butylphenyl)amino-2-bromostyrene 5 were synthesized and polymerized with a palladium-phosphine catalyst to afford the head-to-tail-linked polyradical precursors (1). Oxidation of 1 with the nitrosonium ion solubilized with a crown ether gave the aminium polyradicals (1(+)()) which were durable (half-life > 1 month) at room temperature in air. A high-spin ground state with an average S = (4.5)/2 for 1a(+) was proved even at room temperature by magnetic susceptibility, magnetization, ESR, and NMR measurements.

A SIFT study of the reactions of H 2ONO + ions with several types of organic molecules

A selected ion flow tube (SIFT) study has been carried out of the reactions of hydrated nitrosonium ions, NO +H 2O, which theory has equated to protonated nitrous acid ions, H 2ONO +. One objective of this study was to investigate if this ion exhibits the properties of both a cluster ion and a protonated acid in their reactions with a variety of organic molecules. The chosen reactant molecules comprise two each of the following types—amines, terpenes, aromatic hydrocarbons, esters, carboxylic acids, ketones, aldehydes and alcohols. The reactant H 2ONO + (NO +H 2O) ions are formed in a discharge ion source and injected into helium carrier gas where they are partially vibrationally excited and partially dissociated to NO + ions. Hence, the reactions of the H 2ONO + ions had to be studies simultaneously with NO + ions, the reactions of the latter ions readily being studied by selectively injecting NO + ions into the carrier gas. The results of this study indicate that the H 2ONO + ions undergo a wide variety of reaction processes that depend on the properties of the reactant molecules such as their ionisation energies and proton affinities. These processes include charge transfer with compounds, M, that have low ionisation energies (producing M +), proton transfer with compounds possessing large proton affinities (MH +), hydride ion transfer (MH +), alkyl radical (MR +), alkoxide radical transfer (MOR +), ion–molecule association (NO +H 2OM) and ligand switching (NO +M), producing the ions given in parentheses.

Low‐Temperature Oxidation of Nitrided Iron Surfaces.

AbstractFor Abstract see ChemInform Abstract in Full Text.

Low-Temperature Oxidation of Nitrided Iron Surfaces

The low-temperature (<150 K) oxidation of nitrided iron surfaces exposed to oxygen and water was probed using X-ray photoelectron spectroscopy (XPS), reflection absorption infrared spectroscopy (RAIRS), and mass spectrometry (MS). During exposure of nitrided iron surfaces to oxygen, iron oxynitride (FexNyOz), nitrosonium ions (NO+), as well as nitrite/nitrito- and nitrate-type species were observed. The production of nitrite/nitrito and nitrate species is taken as evidence for the existence of oxygen insertion chemistry into the iron nitride lattice under these low-temperature oxidation conditions. No molecular nitrogen was produced during reactions with oxygen or water in contrast to oxidation studies on other transition metal nitrides. Upon annealing the oxidized overlayer, nitrogen desorbs exclusively as nitric oxide (NO) between 250 and 400 K. In contrast to oxygen, the reactivity of nitrided iron surfaces toward water was limited to the production of adsorbed N−O species.

Role of nitric oxide in cellular iron metabolism.

Iron regulatory proteins (IRP1 and IRP2) control the synthesis of transferrin receptors (TfR) and ferritin by binding to iron-responsive elements (IREs) which are located in the 3' untranslated region (UTR) and the 5' UTR of their respective mRNAs. Cellular iron levels affect binding of IRPs to IREs and consequently expression of TfR and ferritin. Moreover, NO*, a redox species of nitric oxide that interacts primarily with iron, can activate IRP1 RNA-binding activity resulting in an increase in TfR mRNA levels. We have shown that treatment of RAW 264.7 cells (a murine macrophage cell line) with NO+ (nitrosonium ion, which causes S-nitrosylation of thiol groups) resulted in a rapid decrease in RNA-binding of IRP2, followed by IRP2 degradation, and these changes were associated with a decrease in TfR mRNA levels. Moreover, we demonstrated that stimulation of RAW 264.7 cells with lipopolysaccharide (LPS) and interferon-gamma (IFN-gamma) increased IRP1 binding activity, whereas RNA-binding of IRP2 decreased and was followed by a degradation of this protein. Furthermore, the decrease of IRP2 binding/protein levels was associated with a decrease in TfR mRNA levels in LPS/IFN-gamma-treated cells, and these changes were prevented by inhibitors of inducible nitric oxide synthase. These results suggest that NO+-mediated degradation of IRP2 plays a major role in iron metabolism during inflammation.

The reactivity of nitrosyl ruthenium complexes containing polypyridyl ligands

A series of cis nitrosyl complexes containing polypyridyl ligands were prepared and characterized as cis-[RuL(bpy)2(NO)](PF6)3 (L = pyridine, 4-picoline, or 4-acetylpyridine), by elemental analysis, u.v.–vis. and i.r. spectroscopy, and by electrochemical techniques such as cyclic voltammetry, differential pulse voltammetry, spectroelectrochemistry, and coulometry. The complexes exhibit stretching frequencies ν(NO) at ca. 1950 cm−1 indicating that nitrosyl group has a sufficiently high degree of nitrosonium ion (NO+) character. In non-aqueous solution, the reduction of these complexes induce nitrosyl to nitro conversion. In aqueous solution the reduction product is cis-[RuL(bpy)2(NH3)]2+ formed by a six electron mechanism. The nitrosyl compounds are susceptible to nucleophilic attack by hydroxide ion. The equilibrium constants were determined.

Nonendothelial source of nitric oxide in arterioles but not in venules: alternative source revealed in vivo by diaminofluorescein microfluorography.

This study aimed to examine topographic distribution of microvascular NO generation in vivo. To this end, nitrosonium ion (NO+)-sensitive diaminofluorescein diacetate was superfused continuously on the rat mesentery and the fluorescence was visualized in the microvessels through laser confocal microfluorography. Two major sites exhibited a time-dependent elevation of the fluorescence: microvascular endothelia and mast cells. As judged by the fluorescence sensitivity to local application of different inhibitors of NO synthase (NOS), NO availability in arteriolar endothelium and mast cells appeared to be maintained mainly by NOS1, whereas that in venular endothelium greatly depends on NOS3. In venules, the magnitude of inhibitory responses elicited by the inhibitors was positively correlated with the density of leukocyte adhesion. NOS inhibitors significantly reduced, but did not eliminate, the NO+-associated fluorescence in arterioles, capillaries, and venules, suggesting alternative sources of NO in circulation for these microvessels. Immunohistochemistry for NOS isozymes revealed that NOS1 occurred not only in nerve fibers innervated to arterioles but also abundantly in mast cells. Laser flow cytometry of peritoneal cells in vitro revealed abundant expression of NOS1 in mast cells. Interestingly, NOS3 occurred in endothelia of capillaries and venules but not in those of distal arterioles with comparable diameters. These results suggest that the arterioles receive NO from nonendothelial origins involving NOS1 present in nerve terminals and mast cells, whereas venules depend on the endothelial NOS as a major source. Furthermore, nonenzymatic sources of NO from circulating reservoirs constitute a notable fraction throughout different classes of microvessels. The full text of this article is available at http://www.circresaha.org.

Nitric Oxide-Mediated Modulation of Iron Regulatory Proteins: Implication for Cellular Iron Homeostasis

ABSTRACT Iron regulatory proteins (IRP1 and IRP2) control the synthesis of transferrin receptors (TfR) and ferritin by binding to iron-responsive elements (IREs) that are located in the 3′ untranslated region (UTR) and the 5′ UTR of their respective mRNAs. Cellular iron levels affect binding of IRPs to IREs and consequently expression of TfR and ferritin. Moreover, NO •, a redox species of nitric oxide that interacts primarily with iron, can activate IRP1 RNA-binding activity resulting in an increase in TfR mRNA levels and a decrease in ferritin synthesis. We have shown that treatment of RAW 264.7 cells (a murine macrophage cell line) with NO + (nitrosonium ion, which causes S-nitrosylation of thiol groups) resulted in a rapid decrease in RNA-binding of IRP2, followed by IRP2 degradation, and these changes were associated with a decrease in TfR mRNA levels and a dramatic increase in ferritin synthesis. Moreover, we demonstrated that stimulation of RAW 264.7 cells with lipopolysaccharide (LPS) and interferon-γ (IFN-γ) increased IRP1 binding activity, whereas RNA-binding of IRP2 decreased and was followed by a degradation of this protein. Furthermore, the decrease of IRP2 binding/protein levels was associated with a decrease in TfR mRNA levels and an increase in ferritin synthesis in LPS/IFN-γ-treated cells, and these changes were prevented by inhibitors of inducible nitric oxide synthase. These results suggest that NO +-mediated degradation of IRP2 plays a major role in iron metabolism during inflammation.

Chemical ionization of amino and hydroxy group containing arylalkyl compounds with ions in a nitromethane plasma

The gas-phase ion/molecule reactions of organic molecules containing several functional groups, including amino, hydroxy and carboxy groups, have been investigated under nitromethane chemical ionization conditions. Three main reaction channels are observed in the ion source: (a) proton transfer, (b) electron transfer and (c) hydride abstraction. The product ion [M+NO] + is also formed, but in a very low abundance. Initial electrophilic attack of the nitrosonium ion on the aromatic ring is postulated to explain the elimination of HNO from the [M+NO] + adduct ions, observed for all substrates studied. Elimination of water is a characteristic fragmentation pathway for all substrates possessing a benzylic hydroxy group. Fragment ions resulting from cleavage of the molecular ions of the amines, formed by charge transfer, react with the neutral molecules forming two types of adduct ions: [M+immonium] + and [M+C 7H 7] +, which have been characterized through the study of their unimolecular decompositions. The latter provide strong evidence for the existence of two types of structures: a covalent and an ion/molecule complex, that is a non-covalent structure.

The bonding of nitrosonium ion to sulfur in models of cytochrome c-type biomolecules: an ab initio computational study

The possibility that NO+ could bind to one of the two thioether sulfur atoms that covalently link the heme group of cytochrome c to the polypeptide chain, and that the resulting nitrososulfonium ion might undergo S–C cleavage to form a carbocation and a nitrosothiol were investigated computationally, by HF/3-21G(∗) geometry optimizations. The molecules studied were relatively small cytochrome model compounds with only the iron(II) porphyrin unit, and histidine and Me2S molecules coordinated as axial ligands to the heme iron, and R–S+(NO)–CHMe–porphyrin (R=cysteine–histidine, 1; n-butyl, 2; n-pentyl, 3; methyl, 4Me; n-propyl, 4Pr; CH2CH(NH2)CO–NH–CH(Me)–COOH, 5; and 1 without the iron and the Me2S, 6. Molecules 1, 2, 3 and 5 were stationary points on the potential energy surface (they exist at the HF/3-21G(∗) level); on attempted optimization of 4 and 6 the S–CHMe bond breaks tending to give a nitrosothiol RSNO and a delocalized cation +CHMe–porphyrin. This behavior was rationalized by idealizing the iron porphyrin moiety as Fe++ and porphyrin−−; electron density on the porphyrin ring, when not drained away into the iron, can assist with expulsion of the RSNO leaving group (equivalently, departure of RSNO gives a highly delocalized cation +CHMe–porphyrin). Besides iron, a minimum size for R (n-C4H9 or bigger) is needed to hold R in place, perhaps as a manifestation of the ponderal effect. From calculations on smaller molecules, rough CBS-4 water solvation-corrected reaction energies were estimated for loss of NO+ from 1, 2, and 3 (171, 154 and 154kJmol−1, respectively; binding is thermodynamically favored), and for S–CHMe bond breaking of 2 and 3 (−63 and −63kJmol−1, respectively; cleavage is thermodynamically favored).

Formation of nitrosothiols from gaseous nitric oxide at pH 7.4.

Nitric oxide (NO) is generated in biological systems and plays important roles as a regulatory molecule. Its ability to bind to haem iron is well known. Moreover, it may lose an electron, forming the nitrosonium ion, involved in the synthesis of S-nitrosothiols (SNOs). It has been suggested that S-nitrosohaemoglobin (-SNO Hb) and low molecular weight SNOs may act as reservoirs of NO. SNOs are formed in vitro, at strongly acidic pH values; however, the mechanism of their formation at neutral pH values is still debated. In this paper we report the anaerobic formation of SNOs (both high- and low-molecular weight) from low concentrations of NO at pH 7.4, provided Hb is also present. We propose a reaction mechanism entailing the participation of Fehaem in the formation of NO(+) and the transfer of NO(+) either to Cysbeta(93) of Hb or to glutathione; we show that this reaction also occurs in human RBCs.

Structural features of aliphatic N-nitrosamines of 7-azabicyclo[2.2.1]heptanes that facilitate N-NO bond cleavage.

N-Nitrosamines can be considered as potential nitric oxide (NO)/nitrosonium ion (NO(+)) donors. However, the relation of the structures of N-nitrosamines, in particular of aliphatic N-nitrosamines, to the characteristics of release of NO or NO(+) remains unclear. Here we show that aliphatic N-nitrosoamines of 7-azabicyclo[2.2.1]heptanes can undergo heterolytic N-NO bond cleavage. On the basis of the observation of reduced rotational barriers of the N-NO bonds in solution and nitrogen-pyramidal structures of the N-nitroso group in the solid state, we postulate that N-NO bond cleavage of N-nitrosamines is enhanced by a reduction of the resonance in the N-NO group. Computational studies suggest that these structural features of the N-nitrosamines of 7-azabicyclo[2.2.1]heptane are derived from angle strain imposed on the CNC angles.

Nitrosation of melatonin by nitric oxide: a computational study.

Melatonin is being increasingly promoted as a therapeutic agent for the treatment of jet lag and insomnia, and is an efficient free radical scavenger. We have recently characterized a product for the reaction of melatonin with nitric oxide (NO), N-nitrosomelatonin. In the present work, reaction pathways with N1, C2, C4, C6 and C7 as possible targets for its reaction with NO that yield the respective nitroso derivatives have been investigated using semiempirical AM1 computational tools, both in vacuo and aqueous solution. Specifically, two different pathways were studied: a radical mechanism involving the hydrogen atom abstraction to yield a neutral radical followed by NO addition, and an ionic mechanism involving addition of nitrosonium ion to the indolic moiety. Our results show that the indolic nitrogen is the most probable site for nitrosation by the radical mechanism, whereas different targets are probable considering the ionic pathway. These results are in good agreement with previous experimental findings and provide a coherent picture for the interaction of melatonin with NO.

Nitric oxide synthase-I containing cortical interneurons co-express antioxidative enzymes and anti-apoptotic Bcl-2 following focal ischemia: evidence for direct and indirect mechanisms towards their resistance to neuropathology

Neuronal nitric oxide-I is constitutively expressed in ≈2% of cortical interneurons and is co-localized with γ-amino butric acid, somatostatin or neuropeptide Y. These interneurons additionally express high amounts of glutamate receptors which mediate the glutamate-induced hyperexcitation following cerebral injury, under these conditions nitric oxide production increases contributing to a potentiation of oxidative stress. However, perilesional nitric oxide synthase-I containing neurons are known to be resistant to ischemic and excitotoxic injury. In vitro studies show that nitrosonium and nitroxyl ions inactivate N-methyl- d-aspartate receptors, resulting in neuroprotection. The question remains of how these cells are protected against their own high intracellular nitric oxide production after activation. In this study, we investigated immunocytochemically nitric oxide synthase-I containing cortical neurons in rats after unilateral, cortical photothrombosis. In this model of focal ischemia, perilesional, constitutively nitric oxide synthase-I containing neurons survived and co-expressed antioxidative enzymes, such as manganese- and copper-zinc-dependent superoxide dismutases, heme oxygenase-2 and cytosolic glutathione peroxidase. This enhanced antioxidant expression was accompanied by a strong perinuclear presence of the antiapoptotic Bcl-2 protein. No colocalization was detectable with upregulated heme oxygenase-1 in glia and the superoxide and prostaglandin G 2-producing cyclooxygenase-2 in neurons. These results suggest that nitric oxide synthase-I containing interneurons are protected against intracellular oxidative damage and apoptosis by Bcl-2 and several potent antioxidative enzymes. Since nitric oxide synthase-I positive neurons do not express superoxide-producing enzymes such as cyclooxygenase-1, xanthine oxidase and cyclooxygenase-2 in response to injury, this may additionally contribute to their resistance by reducing their internal peroxynitrite, H 2O 2-formation and caspase activation.

A Catalytic Deprotection ofS,S-,S,O- andO,O-Acetals Using Bi(NO3)3⋅5 H2O under Air

S,S-Acetals are smoothly deprotected with air in the presence of a catalytic amount of Bi(NO3)3⋅5 H2O (1–50 mol %) under ambient conditions to regenerate the original carbonyl compounds in good to excellent yield. This mild, simple, and environmentally benign system is successfully applied to the deprotection of S,O- and O,O-acetals and is compatible with various functional groups. From the mechanistic study of the reaction, the catalytic cycle is considered to be composed of the following four steps: (1) the nitrososulfonium ion of the S,S-acetal is formed by attack of nitrosonium ion (NO +) generated from Bi(NO3)3⋅5 H2O through the equilibrium with NO2, (2) the nitrososulfonium ion is hydrolyzed to afford the hemithioacetal and thionitrite, (3) the hemithioacetal collapses to the original carbonyl compound and thiol, which is oxidized by NO + to give disulfide and NO via thionitrite, and (4) the NO captures molecular oxygen from air to regenerate NO2.

Role of calcium in nitric oxide-induced cytotoxicity: EGTA protects mouse oligodendrocytes

Active nitrogen species are overproduced in inflammatory brain lesions in multiple sclerosis (MS) and experimental allergic encephalomyelitis (EAE). NO has been shown to mediate the death of oligodendrocytes (OLs), a primary target of damage in MS. To develop strategies to protect OLs, we examined the mechanisms of cytotoxicity of two NO donors, S-nitroso-N-acetyl-penicillamine (SNAP) and sodium nitroprusside (SNP) on mature mouse OLs. Nitrosonium ion (NO+) rather than NO. mediates damage with both SNAP and SNP, as shown by significant protection with hemoglobin (HbO2), but not with the NO. scavenger PTIO. SNAP and SNP differ in time course and mechanisms of killing OLs. With SNAP, OL death is delayed for at least 6 hr, but with SNP, OL death is continuous over 18 hr with no delay. Relative to NO release, SNP is more toxic than SNAP, due to synergism of NO with cyanide released by SNP. SNAP elicits a Ca2+ influx in over half of the OLs within min. Further, OL death due to NO release from SNAP is Ca2+-dependent, because the Ca2+ chelator EGTA protects OLs from killing by SNAP, and also from killing by the NONOates NOC-9 and NOC-18, which spontaneously release NO. SNP does not elicit a Ca2+ influx, and EGTA is not protective. In comparison to the N20.1 OL cell line (Boullerne et al., [1999] J. Neurochem. 72:1050-1060), mature OLs are (1) more sensitive to SNAP, (2) much more resistant to SNP, (3) sensitive to cyanide, but not iron, and (4) exhibit a Ca2+ influx and EGTA protection in response to NO generated by SNAP.

Determination of S-Nitrosohemoglobin Using a Solid-State Amperometric Sensor

Nitric oxide (NO, nitrogen monoxide), generated in biological systems, plays important roles as a regulatory molecule. Its ability to bind to hemoglobin (Hb) iron is well known. Moreover, it may lose an electron, forming the nitrosonium ion, involved in the synthesis of nitrosothiols (RSNO). It has been suggested that S-nitrosohemoglobin (SNO-Hb) may act as a reservoir of NO. The S-nitrosylation of Hb can be detected after the incubation of CysNO and Hb for 60 min with a molecular ratio (CysNO/hem) of 1:1. Upon increasing the ratio to 10:1, about 50% of total Hb (100% of β-chain −SH 93) was derivatized in 60 min. In this paper, we describe a new method for the quantitative assay of SNO-Hb, after the liberation of NO by Cu2+/Cu+ and the simultaneous assessment of NO by solid-state amperometric sensor. The assay described by us is sensitive, rapid, easy to perform, and inexpensive. For this reason, we believe that it may represent an important analytical improvement for the study of the S-transnitrosylation reactions between RSNO and the Hb Cys-β93 and SNO-Hb and glutathione.

Bromide-free TEMPO-mediated oxidation of primary alcohol groups in starch and methyl α- d-glucopyranoside

TEMPO (2,2,6,6-tetramethylpiperidine-1-oxyl)-mediated oxidation of potato starch and methyl α- d-glucopyranoside (MGP) was performed in the absence of sodium bromide (NaBr) as co-catalyst, solely using sodium hypochlorite (NaOCl) as the primary oxidant. The low reaction rate associated with a bromide-free process was increased by performing the oxidation at increased temperatures. The reaction proceeded stoichiometrically and with high selectivity and with only minor depolymerisation, provided that temperature and pH were kept ≤20 °C and <9.0, respectively. At 20 °C and pH 8.5, the reaction rate was comparable to that of a corresponding oxidation catalysed by NaBr at 2 °C. Consequently, this is a simple approach to raise the TEMPO/NaOCl reaction rate under bromide-free conditions while still maintaining good product properties. At higher oxidation temperatures (≥25 °C) and under more alkaline conditions (pH≥9.0) degradation of the starch skeleton occurred. Simultaneously, side-reactions of the nitrosonium ion lowered the yield of the oxidation. Despite the absence of the NaBr catalyst, the reaction rate-controlling step was found to be the oxidation of the primary hydroxyl groups with the nitrosonium ion. The reaction was first-order in MGP and in TEMPO.

Assay for the transbilayer distribution of glycolipids: selective oxidation of glucosylceramide to glucuronylceramide by TEMPO nitroxyl radicals

In the present study, 2,2,6,6-tetramethylpiperidinooxy nitroxide (TEMPO) has been applied successfully to discriminate between glucosylceramide in the outer and inner leaflets of closed membrane bilayers. The nitroxyl radicals TEMPO and carboxy-TEMPO, once oxidized to nitrosonium ions, are capable of oxidizing residues that contain primary hydroxyl and amino groups. When applied to radiolabeled glucosylceramide in liposomes, oxidation with TEMPO led to an oxidized product that was easily separated from the original lipid by thin-layer chromatography, and that was identified by mass spectrometric analysis as the corresponding acid glucuronylceramide. To test whether oxidation was confined to the external leaflet, TEMPO was applied to large unilamellar vesicles (LUVs) consisting of egg phosphatidylcholine- egg phosphatidylethanolamine;-cholesterol 55:5:40 (mol/mol). TEMPO oxidized most radiolabeled phosphatidylethanolamine, whereas carboxy-TEMPO oxidized only half. Hydrolysis by phospholipase A(2) confirmed that 50% of the phosphatidylethanolamine was accessible in the external bilayer leaflet, suggesting that TEMPO penetrated the lipid bilayer and carboxy-TEMPO did not. When applied to LUVs containing <1 mol% radiolabeled glucosylceramide or short-chain C(6)-glucosylceramide, carboxy-TEMPO oxidized half the glucosylceramide. However, if surface C(6)-glucosylceramide was first depleted by bovine serum albumin (BSA) (extracting 49 +/- 1%), 94% of the remaining C(6)-glucosylceramide was resistant to oxidation. Carboxy-TEMPO oxidized glucosylceramide on the surface of LUVs without affecting inner leaflet glucosylceramide. At pH 9.5 and at 0 degrees C, the reaction reached completion by 20 min.