Spectroscopic studies of nitric oxide (NO) interactions with cobalamins: reaction of NO with superoxocobalamin(III) likely accounts for cobalamin reversal of the biological effects of NO

Recent reports indicate that oxidized cobalamin, Cbl(III), can interfere with the biological effects of nitric oxide (NO) on vascular and visceral smooth muscle and in other systems. In attempting to elucidate the mechanism of these effects of Cbl(III), we reported that a Cbl(III)NO complex could be detected by electron paramagnetic resonance (EPR) spectroscopy, but not by ultraviolet/visible spectroscopy. Subsequently, others concluded that the alleged Cbl(III)NO complex is detectable by ultraviolet/visible, but not by EPR spectroscopy and provided ultraviolet/visible evidence for an alleged Cbl(III)NO complex. We report further investigation of the interaction of NO with Cbl, using both techniques, Fourier transform infrared (FTIR) spectroscopy and mass spectrometry. Our EPR results and the UV/VIS results of others appear to be experimental artifacts that can now, at least in part, be explained. Under conditions where FTIR measurements readily detect a N-O stretching frequency of NO bound to Fe(II), we do not detect a similar signal that can be ascribed to either Cbl(III)NO or Cbl(II)NO, indicating that neither Cbl(III) nor Cbl(II) form a stable complex with NO. Loss of the Cbl(II) EPR signal and mass spectral detection of N2O upon addition of NO to Cbl(II) solutions, demonstrates that Cbl(II), which is present in aerobic Cbl(III) solutions, reduces NO; however, this reaction does not appear to be fast enough to account for the observed biological effects in aerated media. Nitric oxide also reacts rapidly and irreversibly with the superoxo complex of Cbl(III), Cbl(III)O2-, which is always present in aerated solutions of Cbl(III). We believe that this latter reaction accounts for the observed inactivation of NO by Cbl(III) in biological systems. Because Cbl(III)O2- is spontaneously regenerated from Cbl(II) and O2 in aerated solutions, this may constitute a cyclic mechanism for the rapid elimination (oxidation) of NO. Thus, several physicochemical techniques fail to provide convincing evidence for the existence of stable Cbl(III)NO or Cbl(II)NO complexes but do provide evidence that Cbl species participate in redox reactions with NO under aerobic conditions, thereby inhibiting its physiological roles.     

Effects of reactive oxygen and nitrogen species induced by ammonium dinitramide decomposition in aqueous solutions of deoxyribose nucleic acid

Ammonium dinitramide (ADN), a potential rocket fuel, decomposes in water forming NO2. The chemistry of this ADN-released NO2 in oxygenated biological systems is complex both in the number of potential chemical species and in the number of parallel and consecutive reactions that can theoretically occur. High-pressure liquid chromatography (HPLC) studies revealed ADN fragmented deoxyribose nucleic acid (DNA). Damage to DNA standard solutions was caused by at least two major pathways, one arising from reactions of NO2 with oxygen and one arising from a reaction with superoxide (O2-.). The radical species generated when ADN is incubated with standard solutions of DNA, pH 7.5, in the presence of the spin trap agent n-tert-butyl-alpha-nitrone (PBN) was compared with the PBN-radical adducts generated in the presence of ADN and O2-. or of ADN and hydrogen peroxide (H2O2). The ADN-induced PBN radical adducts increased linearly over the 90-minute study period. The values of peak intensity in the presence of O2-. and in the presence of H2O2, were 828% and 7.08%, respectively, of the ADN-induced radicals alone. The synergistic effect of ADN with O2- may provide an understanding of the sensitivity of the rat blastocyst to aDN at the preimplantation stage of development and the lack of toxicity in in vivo studies in tissues high in catalase.     

Chemical biology of nitric oxide: Insights into regulatory, cytotoxic, and cytoprotective mechanisms of nitric oxide

There has been confusion as to what role(s) nitric oxide (NO) has in different physiological and pathophysiological mechanisms. Some studies imply that NO has cytotoxic properties and is the genesis of numerous diseases and degenerative states, whereas other reports suggest that NO prevents injurious conditions from developing and promotes events which return tissue to homeostasis. The primary determinant(s) of how NO affects biological systems centers on its chemistry. The chemistry of NO in biological systems is extensive and complex. To simplify this discussion, we have formulated the "chemical biology of NO" to describe the pertinent chemical reactions under specific biological conditions. The chemical biology of NO is divided into two major categories, direct and indirect. Direct effects are defined as those reactions fast enough to occur between NO and specific biological molecules. Indirect effects do not involve NO, but rather are mediated by reactive nitrogen oxide species (RNOS) formed from the reaction of NO either with oxygen or superoxide. RNOS formed from NO can mediate either nitrosative or oxidative stress. This report discusses various aspects of the chemical biology of NO relating to biological molecules such as guanylate cyclase, cytochrome P450, nitric oxide synthase, catalase, and DNA and explores the potential roles of NO in different biological events. Also, the implications of different chemical reactions of NO with cellular processes such as mitochondrial respiration, metal homeostasis, and lipid metabolism are discussed. Finally, a discussion of the chemical biology of NO in different cytotoxic mechanisms is presented.     

The reaction of NO. with O2.- and HO2.: a pulse radiolysis study

The reactions of NO. with O2.- and with HO2. were studied using the pulse radiolysis technique under pseudo first order conditions where ([O2.-]o + [HO2.]o) > [NO.]o at pH 3.3-10.0. The rate constant of the reaction of NO. with O2.- was determined both by monitoring the decay of O2.- at 250 nm and the formation of ONOO- at 302 nm to be (4.3 +/- 0.5) x 10(9) M-1s-1, independent of ionic strength and pH in the range of 6.1-10.0. The rate constant of the reaction of NO. with HO2.- was determined by following the decay of HO2. at 250 nm to be (3.2 +/- 0.3) x 10(9) M-1s-1 at pH 3.3.     

Tyrosine hydroxylase: mechanisms of oxygen radical formation

A new mechanism of oxygen radical formation in dopaminergic neurons is proposed, based on the oxidative mechanism of tyrosine hydroxylase. The cofactor (6R,6S)-5,6,7,8-tetrahydrobiopterin can rearrange in solution which allows an autoxidation reaction producing O2.-, H2O2 and HO.. The combination of tyrosine hydroxylase and the cofactor produces more oxygen radicals than does the autoxidation of the cofactor. This production of oxygen radicals could be damaging to dopaminergic neurons. In the presence of tyrosine, the enzyme produces less radicals than it does in the absence of tyrosine. Mechanisms are proposed for the generation of reactive oxygen species during the autoxidation of the cofactor and during enzymatic catalysis. The generation, by tyrosine hydroxylase, of very small amounts of oxygen radicals over the period of 65 years could contribute to the oxidative stress that causes Parkinson's disease.     

One electron reduction of metmyoglobin and methemoglobin and the reaction of the reduced molecule with oxygen

We have used the pulse radiolysis technique to reduce with solvated electrons (see article) a single Fe(III) site in methemoglobin and metmyoglobin. The reduction process was followed spectrophotometrically and the reactions rate constants were measured: (see article) =6.5 +/- 1-10(10) M-1-S-1. (see article)=2.5 +/- 0.3-10(10) M-1-S-1. Approx. 60% of the (see article) have reacted with the hemin group, and the rest of the (see article) have probably reacted with the globin moiety. We followed the reaction of the reduced proteins to yield the oxyderivatives and measured the rate constants of the oxygenation process k reduced methemoglobin + O2 = 2.6 +/- 0.6-10(7) M-1-S-1 and k myoglobin + O2 = 1.8 +/- 0.2-10(7) M-1-S-1, all the rate constants were measured at pH = 6.8, I = 0.004, T = 22 +/- 2 degrees C. The high rate constant for reduced methemoglobin indicates that one-site-reduced methemoglobin is probably in the R state, as predicted for methemoglobin from X-ray analysis. The spectra of the reduced and oxygenated species were measured under similar conditions at gamma = 450-650 nm. We were able to follow slight changes in the micro-second time scale, these changes were attributed to conformational changes. We were not able to detect any reaction between the radical (see article) and the hemin group (which would result in a complex such as heme O-2). This may be due to kinetic reasons.     

Mechanisms of autoxidation of the oxygen sensor FixL and Aplysia myoglobin: implications for oxygen-binding heme proteins

On exposure to oxygen, ferrous heme is thought to autoxidize via three distinct mechanisms: (i) dissociation of protonated superoxide from oxyheme; (ii) reaction between a noncoordinated oxygen molecule and pentacoordinate deoxyheme, and (iii) reaction between a noncoordinated oxygen molecule and an intermediate having water coordinated to the ferrous heme iron. The formation of a hexacoordinate aquomet (H2O.Fe3+) species has been proposed to drive mechanism (iii); consequently, heme proteins with a pentacoordinate met (Fe3+) form might be expected to lack this pathway. We have measured the dependence of autoxidation rate on oxygen concentration for Rhizobium meliloti FixL and Aplysia kurodai myoglobin, which have pentacoordinate met forms. For both proteins, the bell shape of this dependence shows that they autoxidize primarily by mechanism (iii), indicating that a hexacoordinate aquomet species is not required for this mechanism. A novel presentation of the oxygen dependence of autoxidation rates that uses heme saturation, rather than oxygen concentration, more clearly reveals the relative contributions of autoxidation pathways.     

Co-capping studies reveal CD8/TCR interactions after capping CD8 beta polypeptides and intracellular associations of CD8 with p56(lck)

A reaction-diffusion model was developed to predict the fate of nitric oxide (NO) released by cells of the immune system. The model was used to analyze data obtained previously using macrophages attached to microcarrier beads suspended in a stirred vessel. Activated macrophages synthesize NO, which is oxidized in the culture medium by molecular oxygen and superoxide (O2-, also released by the cells), yielding mainly nitrite (NO2-) and nitrate (NO3-) as the respective end products. In the analysis the reactor was divided into a "stagnant film" with position-dependent concentrations adjacent to a representative carrier bead and a well-mixed bulk solution. It was found that the concentration of NO was relatively uniform in the film. In contrast, essentially all of the O2- was calculated to be consumed within approximately 2 microm of the cell surfaces, due to its reaction with NO to yield peroxynitrite. The decomposition of peroxynitrite caused its concentration to fall to nearly zero over a distance of approximately 30 microm from the cells. Although the film regions (which had an effective thickness of 63 microm) comprised just 2% of the reactor volume and were predicted to account for only 6% of the NO2- formation under control conditions, they were calculated to be responsible for 99% of the NO3- formation. Superoxide dismutase in the medium (at 3.2 microM) was predicted to lower the ratio of NO3- to NO2- formation rates from near unity to <0.5, in reasonable agreement with the data. The NO3-/NO2- ratio was predicted to vary exponentially with the ratio of O2- to NO release rates from the cells. Recently reported reactions involving CO2 and bicarbonate were found to have important effects on the concentrations of peroxynitrite and nitrous anhydride, two of the compounds that have been implicated in NO cytotoxicity and mutagenesis.     

The effect of low-dose inhalation of nitric oxide in patients with pulmonary fibrosis

The aim of this study was to determine whether low-dose inhalation of nitric oxide (NO) improves pulmonary haemodynamics and gas exchange in patients with stable idiopathic pulmonary fibrosis (IPF). The investigation included 10 IPF patients breathing spontaneously. Haemodynamic and blood gas parameters were measured under the following conditions: 1) breathing room air; 2) during inhalation of 2 parts per million (ppm) NO with room air; 3) whilst breathing O2 alone (1 L.min-1); and 4) during combined inhalation of 2 ppm NO and O2 (1 L.min-1). During inhalation of 2 ppm NO with room air the mean pulmonary arterial pressure (Ppa 25 +/- 3 vs 30 +/- 4 mmHg) and the pulmonary vascular resistance (PVR 529 +/- 80 vs 699 +/- 110 dyn.s.cm-5) were significantly (p < 0.01) lower than levels measured whilst breathing room air alone. However the arterial oxygen tension (Pa,O2) did not improve. The combined inhalation of NO and O2 produced not only a significant (p < 0.01) decrease of Ppa (23 +/- 2 vs 28 +/- 3 mmHg) but also, a remarkable improvement (p < 0.05) in Pa,O2 (14.2 +/- 1.2 vs 11.7 +/- 1.0 kPa) (107 +/- 9 vs 88 +/- 7 mmHg)) as compared with the values observed during the inhalation of O2 alone. These findings suggest that the combined use of nitric oxide and oxygen might constitute an alternative therapeutic approach for treating idiopathic pulmonary fibrosis patients with pulmonary hypertension. However, further studies must first be carried out to demonstrate the beneficial effect of oxygen therapy on pulmonary haemodynamics and prognosis in patients with idiopathic pulmonary fibrosis and to rule out the potential toxicity of inhaled nitric oxide, particularly when used in combination with oxygen.     

Adjuvant radiation therapy for adenocarcinoma of the rectum

The potential for electron transfer quenching of rose bengal triplet (3RB2-) to compete with energy transfer quenching by oxygen was evaluated. Rate constants for oxidative and reductive quenching were measured in buffered aqueous solution, acetonitrile and in small unilamellar liposomes using laser flash photolysis. Biologically relevant quenchers were used that varied widely in structure, reduction potential and charge. Radical ion yields (phi i) were measured by monitoring the absorption of the rose bengal semireduced (RB.3-) and semioxidized (RB.-) radicals. The results in solution were analyzed as a function of the free energy for electron transfer (delta G) calculated using the Weller equation including electrostatic terms. Exothermic oxidative quenching was about 10-fold faster than exothermic reductive quenching in aqueous solution. The quenching rate constants decreased as delta G approached zero in both aqueous and acetonitrile solution. Exceptions to these generalizations were observed that could be rationalized by specific steric or electrostatic effects or by a change in mechanism. The results suggest that electron transfer reactions with some potential quenchers in cells could compete with formation of singlet oxygen [O2(1 delta g)]. Values of phi i were generally greater for reductive quenching and, for oxidative quenching, greater in acetonitrile than in buffer. Electron transfer quenching of 3RB2- in liposomes, below the phase transition temperature was slower than in solution for both lipid-soluble and water-soluble quenchers indicating that these reactions may not compete with formation of O2(1 delta g) during cell photosensitization.     

Accelerated reaction of nitric oxide with O2 within the hydrophobic interior of biological membranes

We demonstrate herein dramatic acceleration of aqueous nitric oxide (NO) reaction with O2 within the hydrophobic region of either phospholipid or biological membranes or detergent micelles and demonstrate that the presence of a distinct hydrophobic phase is required. Per unit volume, at low amounts of hydrophobic phase, the reaction of NO with O2 within the membranes is approximately 300 times more rapid than in the surrounding aqueous medium. In tissue, even though the membrane represents only 3% of the total volume, we calculate that 90% of NO reaction with O2 will occur there. We conclude that biological membranes and other tissue hydrophobic compartments are important sites for disappearance of NO and for formation of NO-derived reactive species and that attenuation of these potentially damaging reactions is an important protective action of lipid-soluble antioxidants such as vitamin E.     

Reactions catalyzed by tetrahydrobiopterin-free nitric oxide synthase

Murine macrophage nitric oxide synthase (NOS) was expressed in E. coli and purified in the presence (holoNOS) or absence (H4B-free NOS) of (6R)-tetrahydro-L-biopterin (H4B). Isolation of active enzyme required the coexpression of calmodulin. Recombinant holoNOS displayed similar spectral characteristics and activity as the enzyme isolated from murine macrophages. H4B-free NOS exhibited a Soret band at approximately 420 nm and, by analytical gel filtration, consisted of a mixture of monomers and dimers. H4B-free NOS catalyzed the oxidation of NG-hydroxy-L-arginine (NHA) with either hydrogen peroxide (H2O2) or NADPH and O2 as substrates. No product formation from arginine was observed under either condition. The amino acid products of NHA oxidation in both the H2O2 and NADPH/O2 reactions were determined to be citrulline and Ndelta-cyanoornithine (CN-orn). Nitrite and nitrate were also formed. Chemiluminescent analysis did not detect the formation of nitric oxide (*NO) in the NADPH/O2 reaction. The initial inorganic product of the NADPH/O2 reaction is proposed to be the nitroxyl anion (NO-) based on the formation of a ferrous nitrosyl complex using the heme domain of soluble guanylate cyclase as a trap, and the formation of a ferrous nitrosyl complex of H4B-free NOS during turnover of NHA and NADPH. NO- is unstable and, under the conditions of the reaction, is oxidized to nitrite and nitrate. At 25 degreesC, the H2O2-supported reaction had a specific activity of 120 +/- 14 nmol min-1 mg-1 and the NADPH-supported reaction had a specific activity of 31 +/- 6 nmol min-1 mg-1 with a KM,app for NHA of 129 +/- 9 microM. HoloNOS catalyzed the H2O2-supported reaction with a specific activity of 815 +/- 30 nmol min-1 mg-1 and the NADPH-dependent reaction to produce *NO and citrulline at 171 +/- 20 nmol min-1 mg-1 with a KM, app for NHA in the NADPH reaction of 36.9 +/- 0.3 microM.     

The effect of absorbed oxygen on the kinetics of chemical reactions on metal surfaces

The effects of adsorbed impurities on the kinetics of reactions on metal surfaces are discussed, in particular the reactions of water vapor and carbon dioxide with iron surfaces are analyzed. The critical oxygen potentials for the onset of ‘poisoning’ of the reactions appear to correspond with the onset of facetting of iron and the formation of surface phases on the iron. The maximum chemical reaction rate constants for the reactions of water vapor and of carbon dioxide on iron surfaces at a given temperature are shown to correspond closely to the chemical reaction rate constants determined from the reduction of iron oxides in their respective systems. The phenomena observed on the reaction of gases with iron and iron oxide surfaces are reported in other metal systems thus the analysis used for the present reactions appears to be applicable to other systems involving metal/gas reactions.     

Identification of drug glucuronides in human urine by RP-HPLC after derivatization

The reactivity of nitric oxide under a given condition is a complex function of its diffusivity and the concentration of reacting partners. Quenching by NO of luminescence from Ru and Pd chelates of mesoporphyrin IX, two molecules which exhibit phosphorescence at room temperature, was utilized to evaluate the gas concentration and apparent diffusion coefficients. The properties of Ru-mesoporphyrin, a dye not previously employed as a probe for O2 or NO, were determined and the assay was verified and used to quantify NO produced by decomposition of nitrosocysteine. The pseudo-second order quenching constants were obtained from Stern-Volmer plots measured under various conditions and used to calculate diffusion coefficients for nitric oxide in solutions, proteins and membranes. The diffusion coefficients were greater at 37 than at 25 degrees C and, at a given temperature, smaller in proteins and membranes than in water. The conclusion is that NO and O2 closely resemble each other in diffusivity but that NO is slightly less lipophilic, resulting in somewhat faster apparent diffusion in protein and slower diffusivity in lipid, relative to O2. Taking a mean diffusion coefficient for NO of 10(-7) cm2s-1, then within 10 s the mean path is 10(-3) cm, or less than the diameter of a single cell. However, at low NO and O2 concentrations, the halflife of NO will be considerably longer than 10 s, and consequently the path of NO diffusion much greater.     

Nitric oxide formation from hydroxylamine by myoglobin and hydrogen peroxide

Hydroxylamine (HA), which is a natural product of mammalian cells, has been shown to possess vasodilatory properties in several model systems. In this study, HA and methyl-substituted hydroxylamines, N-methylhydroxylamine (NMHA) and N,N-dimethylhydroxylamine (NDMHA), have been tested for their ability to generate free diffusible nitric oxide (NO) in the presence of myoglobin (Mb) and hydrogen peroxide. A NO-specific conversion of 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (carboxy-PTIO) to 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl (carboxy-PTI), measured by electron spin resonance (ESR) spectroscopy, along with nitrite and nitrate production, was observed for HA but not for NMHA and NDMHA. ESR measurements at 77 K showed the formation of the ferrous nitrosyl myoglobin, Mb-NO, in the reaction mixtures containing Mb, H2O2 and HA. Our data also demonstrate that Mb-NO is an end product of the reaction pathway involving Mb, H2O2 and HA, rather than a reaction intermediate in the formation of NO. In summary, our results demonstrate a possible pathway of NO formation from HA, however, the significance of this mechanism for bioactivation of HA in vivo is unknown at the present time.     

Efficacy of different schedules in the management of chronic hepatitis C with interferon-alpha

We have evaluated the kinetics of nitrogen dioxide production in a system for inhalation of nitric oxide. In addition to a small fraction of contamination of nitrogen dioxide in the nitric oxide stock gas, a considerable part of the total concentration of nitrogen dioxide is formed immediately after mixing of nitric oxide and oxygen. This initial build-up of nitrogen dioxide is followed by a linear, time-dependent increase in the concentration of nitrogen dioxide. An equation describing the concentration of nitrogen dioxide in the delivery system is formulated: [NO2] = kA x [NO] + kB x [NO]2 x [O2] + kC x t x [NO]2 x [O2], where nitrogen dioxide [NO2] and nitric oxide [NO] concentrations are in parts per million (ppm), oxygen concentration [O2] is expressed as a percentage and contact time (t) is in seconds. The rate constants are kA = 5.12 x 10(-3), kB = 1.41 x 10(-6) and kC = 0.86 x 10(-6). Calculated nitrogen dioxide values correlated well with measured concentrations. This new finding of an initial build-up of nitrogen dioxide has to be taken into consideration if the conversion of nitric oxide to nitrogen dioxide is to be calculated and in the safety guidelines for the use of nitric oxide.     

Cognitive and functional assessments of stroke patients: an analysis of their relation

We have examined the reactions of peroxynitrite with short-chain aliphatic aldehydes to model the reaction of the peroxynitrite anion (ONOO-) with CO2. Aldehydes, like CO2, react rapidly with peroxynitrite and catalyze its decomposition. The pH dependence of the reaction is consistent with the addition of ONOO- (not ONOOH) to the carbonyl carbon atom of the free aldehyde forming a 1-hydroxyalkylperoxynitrite anion adduct (5), which structurally resembles the nitrosoperoxycarbonate adduct (1) formed from the reaction of ONOO- with CO2. Intermediate 5, or the secondary products derived from it, decays to give NO3- and regenerated aldehyde, with small but significant yields of H2O2, organic acids, and organic nitrates. In analogy with the peroxynitrite/CO2 system, it is suggested that 5 undergoes homolytic or heterolytic cleavage at the O-O bond, giving a caged radical pair [RCH(OH)O./ .NO2] (7) or intimate ion pair [RCH(OH)O -/+ NO2] (8). The radicals and ions in intermediates 7 and 8 can recombine within the solvent cage to form 1-hydroxyalkylnitrate [RCH(OH)ONO2] (6), which can then dissociate to give nitrate and regenerate the aldehyde. The aldehyde/ peroxynitrite adducts 5-8 mediate the oxidation of 2,2'-azinobis(3-ethylbenzthiazoline-6-sulfonate) but not the nitration of 4-hydroxyphenylacetate. The significance of these findings is discussed in relation to the mechanism(s) of the CO2-catalyzed isomerization of peroxynitrite to nitrate and biological nitrations involving peroxynitrite/CO2 adducts.     

The kinetics of formation of h2o and co2 during iron oxide reduction

The kinetics of the chemical reaction-controlled reduction of iron oxides by H₂/H₂O and CO/CO₂ gas mixtures are discussed. From an analysis of the systems it is concluded that the decomposition of the oxides takes place by the two dimensional nucleation and lateral growth of oxygen vacancy clusters at the gas/oxide interface. The rates of decomposition of the oxides under conditions of chemical reaction control are dependent not only on the partial pressures of the reacting gases at the reaction temperature but also on the oxygen activity of the prevailing atmosphere. Application of this model to the kinetic data leads to the determination of the maximum chemical reaction rate constants for the decomposition of the iron oxide surfaces. Assuming the reactions H₂ (g) + O(ads) → H₂O(g) andCO(g) + O(ads) → CO₂ (g) to be rate controlling the maximum chemical reaction rate constants for the reduction of iron oxides are given by $$\Phi _{{\text{H}}_{\text{2}} } = 10^{.00} exp \left( {\frac{{ - 69,300}}{{RT}}} \right)mol m^{ - 2} s^{ - 1} atm^{ - 1} $$ and $$\Phi _{CO} = 10^{4.40} \exp \left( {\frac{{103,900}}{{RT}}} \right)mol m^{ - 2} s^{ - 1} atm^{ - 1} $$ The maximum chemical reaction rate constants do not necessarily indicate the maximum rates which can be achieved in practice since these will depend on the limitations imposed by mass transport in the systems. The rate constants are important however since they indicate for the first time the upper limit of any reduction rate in these systems. The fractions of reaction sites which appear to be active on wüstite surfaces in equilibrium with iron are calculated. A direct relationship between chemical reaction rates on liquid iron surfaces and rates on atomically rough iron oxide surfaces is postulated.