Nitrating Agent Research Articles

ChemInform Abstract: Chemistry in Superacids. Part 35. NO2Cl—3MXn Systems: Superelectrophilic Aprotic Nitrating Agents for Deactivated Aromatics.

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Efficient and Selective Mono and Dinitration of Phenols With Cr(NO3)3·2N2O4 as a New Nitrating Agent

Cr(NO3)3·2N2O4 is easily prepared reagent for the efficient and selective mono or dinitration of phenolic compounds. High selectivity ratio of p-nitrophenol formation vs o-nitrophenol is also observed.

XeNO3+: A Gaseous Cation Characterized by a Remarkably Strong Xe−O Bond

The joint application of experimental techniques, such as MIKE, CAD, and FT-ICR spectrometry, and computational methods (DFT and QCISD(T)) has allowed preparation and characterization of gaseous XeNO3+ and XeNO2+ ions. The most stable structure of XeNO3+ has the XeONO2 connectivity, with a relatively short (2.088 Å) Xe−O bond, and represents the gas-phase counterpart of Xe(II) nitrate, previously observed in solution. The XeNO2+ cation is best characterized as a [Xe−NO2]+ complex, with a large (3.018 Å) Xe−NO2 separation and a relatively low dissociation energy, 10.8 kcal mol-1. The reaction of XeNO3+ with selected nucleophiles, including CH3OH, C2H5NO2, CH3COCH3, CH3CN, and C2H5CN, investigated by FT-ICR mass spectrometry, characterizes the ion as a nitrating agent. The unimolecular and collision-induced fragmentation of XeNO3+ and XeNO2+ is discussed.

Vapor phase nitration of benzene over solid acid catalysts (1): Nitration with nitrogen dioxide (NO2)

Vapor phase nitration of benzene over acidic catalysts is expected to be a clean process with no sulfuric acid waste. We investigated this process over acidic catalysts utilizing nitrogen dioxide (NO2) as a nitrating agent, and found that several mixed metal oxides, such as silica-alumina, zinc oxide-titania, and tungsten oxide-molybdenum oxide, exhibited a fairly good activity. Among them, WO3−MoO3 is the most active; however, only this catalyst deviated from the linear relationship between the activity and the acidity. In order to explain this discrepancy, several factors such as acid amount, acid strength, and BET surface area, are examined in detail.

Peroxynitrite: a putative cytotoxin.

In recent years it has become apparent that peroxynitrite, which is one of the toxic metabolites originating from the reaction of nitric oxide and superoxide presents a number of pathologic states in which free radicals are thought to be involved. Peroxynitrite is capable of oxidizing a wide variety of biomolecules including plasma, proteins, lipids, carbohydrates and nucleic acids. Peroxynitrite is involved in the hydroxylation of aromatic compounds and acts as a nitrating agent. It modifies free or protein-associated tyrosine residues to give nitrotyrosines, leaving a marker detectable in vivo. Peroxynitrite has been implicated in the pathophysiology of a variety of diseases including inflammation, atherosclerosis, arthritis, endotoxemia, ischaemia-reperfusion injury, or acute respiratory distress syndrome. Development of specific peroxynitrite scavengers may provide new approaches for the effective treatment of these disease states.

3-Nitrotyrosine in the proteins of human plasma determined by an ELISA method

The modification of tyrosine residues in proteins to 3-nitrotyrosine by peroxynitrite or other potential nitrating agents has been detected in biological systems that are subject to oxidative stress. A convenient semi-quantitative method has been developed to assay nitrated proteins in biological fluids and homogenates using a competitive ELISA developed in our laboratory. This assay selectivity detected 3-nitro-l-tyrosine residues in a variety of peroxynitrite-treated proteins (BSA, human serum albumin (HSA), alpha1-antiprotease inhibitor, pepsinogen and fibrinogen) and also in a nitrated peptide, but had a low affinity for free 3-nitro-L-tyrosine and 3-chloro-L-tyrosine. The IC50 values for the inhibition of antibody binding by different nitrated proteins were in the range 5-100 nM, suggesting that the antibody discriminated between nitrotyrosine residues in different environments. The presence of nitrotyrosine in plasma proteins was detected by Western blot analysis and quantified by the ELISA. A concentration of 0. 12+/-0.01 microM nitro-BSA equivalents was measured in the proteins of normal plasma which was increased in peroxynitrite-treated plasma and was elevated in inflammatory conditions. HSA and low-density lipoprotein (LDL) isolated from plasma contained 0.085+/-0.04 and 0. 03+/-0.006 nmol nitro-BSA equivalents/mg protein, respectively. Comparison of the level of nitration in peroxynitrite-treated HSA and LDL in the presence and absence of plasma indicates that nitration and presumably oxidation is inhibited by plasma antioxidants. The presence of nitrotyrosine in LDL is consistent with previous reports implicating peroxynitrite in the oxidative modification of lipoproteins and the presence of a low concentration of oxidized LDL in the blood.

Electrosynthetic immobilisation of proteins for bioanalysis

Use of electrosynthetic methodology allows the production of hen egg-white lysozyme (HEWL) either mononitrated at tyrosine 23 or bisnitrated at tyrosines 20 and 23, but never nitrated at tyrosine 53. This is a different sequence from that obtained by the chemical nitrating agent tetranitromethane, and when reduced by dithionite, the selectively modified enzyme can be anchored at pH 5 via the unique aromatic amino group to magnetic beads or other suitable matrices. HEWL so immobilised loses less than 10% of cell-wall lytic activity compared with the approximately 50% loss of activity when immobilised by conventional methodology at pH 9 via essentially random reaction at lysine residues and other functionalities which are nucleophilic at this pH. This result offers promise as a general method for selective protein immobilisation in biosensors and similar applications. © 1998 John Wiley & Sons, Ltd.

Electrooxidative nitration of phenols at copper electrodes

p-Cresol, as a small-molecule model for tyrosine residues in proteins, undergoes electrooxidative nitration in the presence of a nitrogen source, for example ammonia, in mildly alkaline aqueous solution at potentials in the range 0.8 to 1.4 V (vs sce). Anodized copper is the best electrode material of those studied and nitrogen sources in the increasing order of effectiveness are amides < amines ≅ proteins < ammonia, the latter giving a total of nitrocresols of ~ 30% from an initial p-cresol concentration of 0.5 mM. Azulene also nitrates in these conditions, but phenylethers (4-methyl methoxybenzene and 1,2-dimethoxy benzene) do not. The protein hen egg-white lysozyme (HEWL), in the absence of any other nitrogenous species, acts as a source of nitrating agent in the electrooxidative nitration of p-cresol thus substantiating our earlier finding of selective tyrosine nitration in proteins in the absence of any other nitrogen source. This electronitration reaction, unique in that there is no NO bond in any initial species, provides a novel and environmentally friendly route in mild conditions and is of particular benefit in the selective covalent modification of proteins.

Effects of carbon dioxide/bicarbonate on induction of DNA single-strand breaks and formation of 8-nitroguanine, 8-oxoguanine and base-propenal mediated by peroxynitrite

Carbon dioxide has been reported to react with peroxynitrite (ONOO −), a strong oxidant and nitrating agent, to form an ONO 2CO 2 − adduct, altering the reactivity characteristic of peroxynitrite. We found that bicarbonate (0–10 mM) caused a dose-dependent increase of up to 6-fold in the formation of 8-nitroguanine in calf-thymus DNA incubated with 0.1 mM peroxynitrite, whereas it produced no apparent effect on 8-oxoguanine formation. In contrast, bicarbonate inhibited peroxynitrite-induced strand breakage in plasmid pBR322 DNA and thymine-propenal formation from thymidine. We conclude that CO 2 HC0 3 − reacts with peroxynitrite to form a potent nitrating agent, but also to inactivate hydroxyl-radical-like activity of peroxynitrous acid.

Interaction of nitrogen monoxide with cytochrome P-450 monitored by surface-enhanced resonance Raman scattering

The reaction of mammalian cytochrome P-450 2B4 with nitrogen monoxide and oxygen has been studied by surface-enhanced resonance Raman scattering (SERRS) to obtain sharp and definitive information in situ on the nature of the changes in the active site pocket. The initial reaction produces a six co-ordinate low spin haem-nitrogen monoxide adduct. A slower reaction leads to the irreversible formation of a five co-ordinate high-spin iron (III) haem with no nitrogen monoxide bound to it and to the nitration of an aromatic side chain, probably a tyrosine, in the proximity of the active site. In the presence of excess nitrogen monoxide, the second reaction is controlled by oxygen concentration. The sequence of events corresponds to the biphasic inhibition induced by NO in other cytochromes P-450 and peroxidases and is postulated to occur by the formation of a nitrating agent at the haem followed by diffusion to the tyrosine. The nitrated amino acid and the oxidation and spin state of the haem are observed easily by SERRS with low concentrations of protein making it a particularly suitable method for the investigation of reactions of NO in complex biological matrices.

A Facile Synthesis of Dinitro[2.2]metacyclophane and Its Partial Reduction

AbstractBoth aromatic rings in [2.2]metacyclophanes(MCPs) can be easily nitrated in good yields by using the HNO3/dichloromethane system under mild conditions. This facile method is clearly superior to the nitration by strong nitrating agents such as nitronium tetrafluoroborate. Partial reduction of dinitro[2.2]MCP (2a) affords the intriguing aminonitro[2.2]MCP (3) which is one of the ultimate models of intramolecular charge‐transfer interaction. Compound 3 shows a significant bathochromic shift due to through‐space interaction between two aromatic rings.

Peroxynitrite-dependent tryptophan nitration.

Peroxynitrite (ONOO-), the reaction product of superoxide (O2.-) and nitric oxide (.NO), nitrates tyrosine and other phenolics. We report herein that tryptophan is also nitrated by peroxynitrite in the absence of transition metals to one predominant isomer of nitrotryptophan, as determined from spectral characteristics and liquid chromatography-mass spectrometry analysis. At high peroxynitrite to tryptophan ratios, other oxidation products were detected as well. The amount of nitrotryptophan formed from peroxynitrite increased at acidic pH, with an apparent pKa of 7.8. High concentrations of Fe(3+)-EDTA were required to enhance peroxynitrite-induced nitrotryptophan formation, while addition of up to 15 microM Cu/Zn superoxide dismutase had a minimal effect on tryptophan nitration. Cysteine, ascorbate, and methionine decreased nitrotryptophan yield to an extent similar to that predicted by their reaction rates with ground-state peroxynitrite, and typical hydroxyl radical scavengers partially inhibited nitration. Plots of the observed rate constant of nitrotryptophan formation vs tryptophan concentration presented downward curvatures. Thus, the kinetics of metal-independent nitration reactions were interpreted in terms of two parallel mechanisms. In the first one, ground-state peroxynitrous acid nitrated tryptophan with a second-order rate constant of 184 +/- 11 M-1 s-1 at 37 degrees C. The activation enthalpy was 9.1 +/- 0.3 kcal mol-1, and the activation entropy was -19 +/- 1 cal mol-1 K-1. In the second mechanism, ONOOH*, an activated intermediate derived from trans-peroxynitrous acid formed in a steady state, was the nitrating agent.

20] Peroxynitrite-dependent tyrosine nitration catalyzed by superoxide dismutase, myeloperoxidase, and horseradish peroxidase

This chapter discusses the peroxynitrite-dependent tyrosine nitration catalyzed by superoxide dismutase, myeloperoxidase, and horseradish peroxidase. Peroxynitrite is a potent nitrating agent formed by the near diffusion limited reaction of superoxide and nitric oxide (NO). Peroxynitrite is more cytotoxic to Escherichia coli and Trypanosoma cruzi than NO, superoxide, or hydrogen peroxide. Peroxynitrite is a strong oxidant capable of yielding the chemistry commonly attributed to hydroxyl radical damage. The peroxynitrite anion can exist in two stable conformers. The cis conformer, which is ring-like because of overlapping of the terminal oxygen orbitals, is slightly more stable and is the only conformer found in alkaline solution, cis -Peroxynitrite cannot directly isomerize to nitrate. The simplest means to observe tyrosine nitration is to mix rapidly a small volume of peroxynitrite with a solution containing the target phenol and measure the resulting yellow absorbance from the nitrophenolic. Because peroxynitrite rapidly decomposes at neutral pH, the secret to reproducibility involves rapid mixing and accurate pipetting.

Quantitation of Protein Tyrosine, 3-Nitrotyrosine, and 3-Aminotyrosine Utilizing HPLC and Intrinsic Ultrviolet Absorbance

Nitrotyrosine has been found in the urine of humans with no known exposure to exogenous nitrating agents. We have shown that peroxynitrite, a nitrating agent formed by the near diffusion-limited reaction of nitric oxide with superoxide, is formed by activated macrophages. Using an antibody which recognizes nitrated tyrosine residues in proteins, we have obtained immunohistological evidence for nitration in a number of pathological conditions amenable to peroxynitrite formation. We have developed an HPLC method as a means of confirming the presence of nitrotyrosine. Quantitation of small amounts of nitrotyrosine in complex protein mixtures presents special problems of sensitivity and specificity which are often exacerbated by traditional amine-derivatization of all amino acids. An HPLC method for proteins hydrolysates is described which relies on intinsic UV absorbance of nitrotyrosine at 280 nm (which is fivefold higher than tyrosine at pH 3.5) for quantitation and on its unique absorbance at 355-365 nm for partial identification. Since only aromatic acids are detected, sample sizes can be increased to permit detection of small amounts of nitrotyrosine without introducing interference. Treatment of BSA with 4mM peroxynitrite resulted in nitration of 12.8% of total tyrosine residues; a small fraction (0.9%) of the total was detected as aminotyrosine. A total of 9.6% of the tyrosine residues in fresh plasma were nitrated as a result of treatment with 4mM peroxytyrosinee during hydrolysis. The advantages of this method over traditional amine-dervatized amino acid analysis are discussed and modifications which would further increase sensitivity to aminotyrosine are presented.

Nitration of nonactivated arenes with a ternary system NO–NO2–O2. Mechanistic implications of the Kyodai-nitration

The ternary mixture NO–NO2–O2 has been found to be more effective than the binary mixture NO2–O2 as a nitrating agent for nonactivated arenes, such as toluene and chlorobenzene; the results may be rationalized in terms of an initial electron transfer mechanism in which nitrogen trioxide, formed from nitrogen monoxide and dioxygen, oxidizes the arene to its radical cation.

Production of nitric oxide and peroxynitrite in the lung during acute endotoxemia.

Nitric oxide is a short-lived cytotoxic mediator that has been implicated in the pathogenesis of endotoxin-induced tissue injury and septic shock. In the present studies we determined whether this mediator is produced in the lung during acute endotoxemia. We found that intravenous injection of rats with bacterially derived lipopolysaccharide (LPS), a condition that induces acute endotoxemia, caused a time-dependent increase in inducible nitric oxide synthase (iNOS) mRNA expression in the lung, which reached a maximum after 24 h. This was correlated with nitric oxide production in the lung as measured by electron paramagnetic spin trapping, which was detectable within 6 h. Alveolar macrophages (AMs) and interstitial macrophages (IMs) isolated from rats 6-12 h after induction of acute endotoxemia were also found to exhibit increased nitric oxide production in response to in vitro stimulation with interferon-gamma (IFN-gamma) and LPS measured by nitrite accumulation in the culture medium. The effects of acute endotoxemia on nitric oxide production by these cells were, however, transient and returned to control levels by 24 h in AMs and 36 h in IMs. Interestingly, although nitrite accumulation in the culture medium of IMs isolated 48 h after induction of acute endotoxemia and stimulated with low concentrations of IFN-gamma and LPS was reduced, when compared with cells from control animals, these cells, as well as AMs, continued to express high levels of iNOS protein and mRNA. This was correlated with increased peroxynitrite production by the cells. Peroxynitrite has been shown to act as a nitrating agent and can generate nitrotyrosine residues in proteins. Using a specific antibody and immunohistochemistry, we found evidence of nitrotyrosine residues in sections of lungs 48 h after treatment of rats with endotoxin. These data suggest that nitric oxide produced by IMs and AMs can react with superoxide anion to form peroxynitrite. Taken together, the present studies demonstrate that AMs and IMs are activated following acute endotoxemia to produce reactive nitrogen intermediates and that both cell types contribute to inflammatory responses in the lung.

Vapor-Phase Aromatic Nitration with Dinitrogen Tetroxide over Solid Acids: Kinetics and Mechanism

The mechanism of the vapour-phase mononitration of aromatics with dinitrogen tetroxide has been investigated over silica-alumina and the large-pore zeolite β (BEA). A Rideal-Eley rate law is observed in both cases, showing that the reaction occurs between the chemisorbed aromatic and the gaseous nitrating agent. The effects of substituents are better correlated by a relation between the activity and the ionisation potential of the aromatic than by the Hammett equation. Both results conflict with the "classical" electrophilic aromatic substitution (SEAr), but agree with the formation of an aromatic radical cation intermediate at the surface of the catalyst and its reaction with a nitrogen dioxide radical in the rate determining step. The generation of electron-acceptor sites on the catalyst surface by nitrogen dioxide radicals is suggested. Addition of water in the feed inhibits the reaction by competitive adsorption over silica-alumina, but improves the efficiency of BEA, certainly by "steam cleaning" of the surface of this hydrophobic catalyst.

Nitro-substituted porphyrazines

This nitration of tetra (tert-butyl)porphyrazine and its metal complexes with different nitrating agents has given their mono-, di-, tri-, and tetra-nistro-derivatives; the metal complexes have also been obtained by the metallation of the corresponding metal—free nitroporphyrazines. The structure of the synthesized compounds has been confirmed by electron and mass spectra, and by PMR spectra.

Microemulsions as Media for Organic Synthesis: Selective Nitration of Phenol to ortho-Nitrophenol Using Dilute Nitric Acid

The paper reports near-selective nitration of phenol to ortho -nitrophenol using a suitably prepared microemulsion solution in the presence of dilute nitric acid. The case example indicates that microemulsions, besides enhancing the reaction rates, can also be selective to give the desired product. Microemulsion solution also affords the use of dilute acid as a nitrating agent.

Facile preparation of α-nitroketones from enol silyl ethers

Abstract Treatment of enol silyl ethers with the mild nitrating agent tetranitromethane gives good yields of α-nitroketones at room temperature and below.