Nitric Oxide (NO)

Nitric oxide or nitrogen monoxide is a chemical compound with chemical formula NO. This gas is an important signaling molecule in the body of mammals, including humans, and is an extremely important intermediate in the chemical industry. It is also an air pollutant produced by cigarette smoke, automobile engines and power plants.

NO is an important messenger molecule involved in many physiological and pathological processes within the mammalian body both beneficial and detrimental. Appropriate levels of NO production are important in protecting an organ such as the liver from ischemic damage. However sustained levels of NO production result in direct tissue toxicity and contribute to the vascular collapse associated with septic shock, whereas chronic expression of NO is associated with various carcinomas and inflammatory conditions including juvenile diabetes, multiple sclerosis, arthritis and ulcerative colitis.

Nitric oxide should not be confused with nitrous oxide (N₂O), a general anaesthetic and greenhouse gas, or with nitrogen dioxide (NO₂) which is another air pollutant. The nitric oxide molecule is a free radical, which is relevant to understanding its high reactivity.

Despite being a simple molecule, NO is a fundamental player in the fields of neuroscience, physiology, and immunology, and was proclaimed “Molecule of the Year” in 1992

Reactions

• When exposed to oxygen, NO is converted into nitrogen dioxide.
  2NO + O₂ → 2NO₂
  This conversion has been speculated as occurring via the ONOONO intermediate. In water, NO reacts with oxygen and water to form HNO₂ or nitrous acid. The reaction is thought to proceed via the following stoichiometry:
  4 NO + O₂ + 2 H₂O → 4 HNO₂
• NO will react with fluorine, chlorine, and bromine to form the XNO species, known as the nitrosyl halides, such as nitrosyl chloride. Nitrosyl iodide can form but is an extremely short lived species and tends to reform I₂.
  2NO + Cl₂ → 2NOCl
• Nitroxyl (HNO) is the reduced form of nitric oxide.

Preparation

• Commercially, NO is produced by the oxidation of ammonia at 750°C to 900°C (normally at 850°C) in the presence of platinum as catalyst:
  4NH₃ + 5O₂ → 4NO + 6H₂O
  The uncatalyzed endothermic reaction of O₂ and N₂ which is performed at high temperature (>2000°C) with lightning has not been developed into a practical commercial synthesis:
  N₂ + O₂ → 2NO
• In the laboratory, it is conveniently generated by reduction of nitric acid:
  8HNO₃ + 3Cu → 3Cu(NO₃)₂ + 4H₂O + 2NO
• or by the reduction of nitrous acid:
  2 NaNO₂ + 2 NaI + 2 H₂SO₄ → I₂ + 4 NaHSO₄ + 2 NO
  2 NaNO₂ + 2 FeSO₄ + 3 H₂SO₄ → Fe₂(SO₄)₃ + 2 NaHSO₄ + 2 H₂O + 2 NO
  3 KNO₂(l) + KNO₃ (l) + Cr₂O₃(s) → 2 K₂CrO₄(s) + 4 NO (g)
  The iron(II) sulfate route is simple and has been used in undergraduate laboratory experiments.
• So-called NONOate compounds are also used for NO generation.

Coordination Chemistry

NO forms complexes with all transition metals to give complexes called metal nitrosyls. The most common bonding mode of NO is the terminal linear type (M-NO). The angle of the M-N-O group can vary from 160-180° but are still termed as "linear". In this case the NO group is formally considered a 3-electron donor. In the case of a bent M-N-O conformation the NO group can be considered a one electron donor. Alternatively, one can view such complexes as derived from NO⁺, which is isoelectronic with CO.

Nitric oxide can serve as a one-electron pseudohalide. In such complexes, the M-N-O group is characterized by an angle between 120-140°.

The NO group can also bridge between metal centers through the nitrogen atom in a variety of geometries.

Measurement of nitric oxide concentration

The concentration of nitric oxide can be determined using a simple chemiluminescent reaction involving ozone: A sample containing nitric oxide is mixed with a large quantity of ozone. The nitric oxide reacts with the ozone to produce oxygen and nitrogen dioxide. This reaction also produces light (chemiluminescence), which can be measured with a photodetector. The amount of light produced is proportional to the amount of nitric oxide in the sample.

NO + O₃ → NO₂ + O₂ + light

Other methods of testing include electroanalysis(amperometric approach), where NO reacts with an electrode to induce a current or voltage change. The detection of NO radicals in biological tissues is particularly difficult due to the short lifetime and concentration of these radicals in tissues. One of the few practical methods is spin trapping of nitric oxide with iron-dithiocarbamate complexes and subsequent detection of the mono-nitrosyl-iron complex with Electron Paramagnetic Resonance (EPR).

A group of fluorescent dye indicators exist that are also available in acetylated form for intracellular measurements. The most common compound is 4,5-diaminofluorescein (DAF-2).

Production environmental effects

From a thermodynamic perspective, NO is unstable with respect to O₂ and N₂, although this conversion is very slow at ambient temperatures in the absence of a catalyst. Because the heat of formation of NO is endothermic, its synthesis from molecular nitrogen and oxygen requires elevated temperatures, >1000°C. A major natural source is lightning. The use of internal combustion engines has drastically increased the presence of nitric oxide in the environment. One purpose of catalytic converters in cars is to minimize NO emission by catalytic reversion to O₂ and N₂. Nitric oxide in the air may convert to nitric acid, which has been implicated in acid rain. Furthermore, both NO and NO₂ participate in ozone layer depletion. Nitric oxide is a small highly diffusible gas and a ubiquitous bioactive molecule.

Technical applications

Although NO has relatively few direct uses, it is produced on a massive scale as an intermediate in the Ostwald process for the synthesis of nitric acid from ammonia. In 2005, the US alone produced 6M metric tons of nitric acid. It finds use in the semiconductor industry for various processes. In one of its applications it is used along with nitrous oxide to form oxynitride gates in CMOS devices.

Miscellaneous applications

Nitric oxide can be used for detecting surface radicals on polymers. Quenching of surface radicals with nitric oxide results in incorporation of nitrogen, which can be quantified by means of X-ray photoelectron spectroscopy.

Biological functions

NO is one of the few gaseous signaling molecules known. It is a key vertebrate biological messenger, playing a role in a variety of biological processes. Nitric oxide, known as the 'endothelium-derived relaxing factor', or 'EDRF', is biosynthesised endogenously from arginine and oxygen by various nitric oxide synthase (NOS) enzymes and by reduction of inorganic nitrate. The endothelium (inner lining) of blood vessels use nitric oxide to signal the surrounding smooth muscle to relax, thus resulting in vasodilation and increasing blood flow. Nitric oxide is highly reactive (having a lifetime of a few seconds), yet diffuses freely across membranes. These attributes make nitric oxide ideal for a transient paracrine (between adjacent cells) and autocrine (within a single cell) signaling molecule. The production of nitric oxide is elevated in populations living at high-altitudes, which helps these people avoid hypoxia by aiding in pulmonary vasculature vasodilation. Effects include vasodilatation, neurotransmission, modulation of the hair cycle, production of reactive nitrogen intermediates and penile erections (through its ability to vasodilate). Nitroglycerin and amyl nitrite serve as vasodilators because they are converted to nitric oxide in the body. Sildenafil, popularly known by the trade name Viagra, stimulates erections primarily by enhancing signaling through the nitric oxide pathway in the penis.

Nitric oxide (NO) contributes to vessel homeostasis by inhibiting vascular smooth muscle contraction and growth, platelet aggregation, and leukocyte adhesion to the endothelium. Humans with atherosclerosis, diabetes or hypertension often show impaired NO pathways. A high-salt intake was demonstrated to attenuate NO production, although bioavailability remains unregulated.

Nitric oxide is also generated by phagocytes (monocytes, macrophages, and neutrophils) as part of the human immune response. Phagocytes are armed with inducible nitric oxide synthase (iNOS) which is activated by interferon-gamma (IFN-γ) as a single signal or by tumor necrosis factor (TNF) along with a second signal. Conversely, transforming growth factor-beta (TGF-ß) provides a strong inhibitory signal to iNOS whereas interleukin-4 (IL-4) and IL-10 provide weak inhibitory signals. In this way the immune system may regulate the armamentarium of phagocytes that play a role in inflammation and immune responses. Nitric oxide secreted as an immune response is as free radicals and is toxic to bacteria; the mechanism for this include DNA damage and degradation of iron sulfur centers into iron ions and iron-nitrosyl compounds. In response, however, many bacterial pathogens have evolved mechanisms for nitric oxide resistance. Because nitric oxide might serve as an inflammometer in conditions like asthma, there has been increasing interest in the use of exhaled nitric oxide as a breath test in diseases with airway inflammation.

Nitric oxide can contribute to reperfusion injury when an excessive amount produced during reperfusion (following a period of ischemia) reacts with superoxide to produce the damaging oxidant peroxynitrite. In contrast, inhaled nitric oxide has been shown to help survival and recovery from paraquat poisoning, which produces lung tissue damaging superoxide and hinders NOS metabolism.

In plants, nitric oxide can be produced by any of four routes: (i)L-arginine-dependent nitric oxide synthase (although the existence animal NOS homologs in plants is debated), (ii) by plasma membrane-bound nitrate reductase, (iii) by mitochondrial electron transport chain, or (iv) by non-enzymatic reactions. It is a signaling molecule, acts mainly against oxidative stress and also plays a role in plant pathogen interactions. Treating cut flowers and other plants with nitric oxide has been shown to lengthen the time before wilting.

A biologically important reaction of nitric oxide is S-nitrosylation, the conversion of thiol groups, including cysteine residues in proteins, to form S-nitrosothiols (RSNOs). S-Nitrosylation is a mechanism for dynamic, post-translational regulation of most or all major classes of protein.

Mechanism of action

There are several mechanisms by which NO has been demonstrated to affect the biology of living cells. These include oxidation of iron containing proteins such as ribonucleotide reductase and aconitase, activation of the soluble guanylate cyclase, ADP ribosylation of proteins, protein sulphhydryl group nitrosylation, and iron regulatory factor activation. NO has been demonstrated to activate NF-κB in peripheral blood mononuclear cells, an important transcription factor in iNOS gene expression in response to inflammation. It was found that NO acts through the stimulation of the soluble guanylate cyclase which is a heterodimeric enzyme with subsequent formation of cyclic GMP. Cyclic GMP activates protein kinase G, which caused phosphorylation (and therefore inactivation) of myosin light-chain kinase and leads ultimately to the dephosphorylation of the myosin light chain, causing smooth muscle relaxation.

Use in pediatric intensive care

Nitric oxide/oxygen blends are used in critical care to promote capillary and pulmonary dilation to treat primary pulmonary hypertension in neonatal patients post meconium aspiration and related to birth defects. These are often a last-resort gas mixture before the use of ECMO. Nitric oxide therapy has the potential to significantly increase the quality of life and in some cases save the lives of infants at risk for pulmonary vascular disease.

Nutraceutical marketing

GNC has begun to sell an oral "nitric oxide" product targeted for bodybuilders, with the claim that it dramatically increases muscle growth. The claim is grounded in an understanding of NO as being a vasodilator, and when taken prior to and after workouts, it enables muscles to receive more blood and therefore, more oxygen and nutrients. This is critical to maximal muscle exertion during training and recovery afterward. However, there are currently no valid studies supporting the hypothesis that orally ingested NO actually will cause vasodilation; additionally, while users of some supplements have claimed to experience results, these results are generally attributable to ingredients besides NO itself (proteins, creatine etc).

Effects of Various NO Levels