Azide Complexes Research Articles

Simple Purification and Improvement of Stability of Alcohol Oxidase from a Methanol Yeast,Candida boidiniiS2, by Forming an Enzyme–Azide Complex

It was found that azide bound to alcohol oxidase non-covalently and caused a color change from yellow to red. Alcohol oxidase was purified with a high yield from methanol-limited chemostat-grown cells of Candida boidinii S2 as an enzyme-azide complex by a simple procedure. That is, a cell-free extract was prepared from cells treated with a cationic detergent, Cation M2, and alcohol oxidase amounted to more than 80% of the soluble protein. Inactivation of the enzyme by H2O2 and aldehydes was decreased in the complex. The results were discussed as to the production of various aldehydes with the yeast cells.

Studies on ruthenium(II) macrocyclic amine complexes. Synthesis and X-ray structure of the (R,S,R,R) isomer of trans-[Ru II (L)(N3)(NCMe)]PF6(L = 1,4,8,11-tetramethyl-1,4,8,11-tetra-azacyclotetradecane)

The reaction between K2[RuCl5(NO)] and L (L = 1,4,8,11-tetramethyl-1,4,8,11-tetraazacyclotetradecane) in ethanol at reflux gave an intermediate product which, on treatment with N2H4·H2O in acetonitrile yielded trans-[RuII(L)(N3)(NCMe)]PF6. This azide complex crystallizes in space group P21/n, with a= 14.775(3), b= 18.083(2), c= 18.167(2)A, β= 90.26(1)°, and Z= 8. One of the two independent cations has a disordered ligand L, and the methyl groups in L of the ordered cation adopt a ‘three-up, one-down’(R,S,R,R) configuration. The measured Ru–N(acetonitrile) bond lengths, 1.976(7) and 2.003(7)A, are significantly shorter than the corresponding distance of 2.149(6)A in the (R,S,R,R) isomer of trans-[Ru(L)O(NCMe)][PF6]2, attesting to a greater trans influence of the oxo ligand as compared to the azide ligand.

Kinetics of 1,3-dipolar cycloadditions of dimethylacetylenedicarboxylate to LCo(AcAc) 2N 3

A series of LCo(AcAc) 2N 3 complexes (where AcAc is the monoanion of 2,4-pentanedione and L is ( p-ClC 6H 4) 3P, (C 6H 5) 3P, ( p-CH 3C 6H 4) 3P, ( p- CH 3OC 6H 4) 3P, 1-phenyldibenzophosphole or pyridine) have been prepared and characterized by infrared, 1H and 13C{ 1H} NMR spectroscopy, cyclic voltammetry and elemental analyses. All the phosphine complexes are cis and the pyridine complexes have been isolated as both cis and trans isomers. In solution the pyridine complexes exist as an equilibrium mixture of the two isomers with the cis isomer predominating (k eq = 2.49). The isomerization process is relatively slow; t = 40 min for approach to equilibrium. These complexes undergo 1,3-dipolar cycloaddition reactions with dimethylacetylenedi- carboxylate (DAD) to produce trans isomers of LCo- (AcAc) 2X where X is the N(2)-bound 4,5-bis(methoxycarbonyl)-1,2,3-triazolate. The kinetics of these reactions have been investigated by infrared spectroscopy. For the PyCo(AcAc) 2N 3 complexes the cyclo- addition reactions are associative, second-order processes with rates that are independent of the geometry of the complex. The cycloaddition reactions of the R 3PCo(AcAc) 2N 3 complexes are first- order processes with rates that are independent of the DAD concentration. For the ( p-x-C 6H 5) 3PCo- (AcAc) 2N 3 complexes, the rates of the cycloaddition reactions are linearly related to the Hammet σ constants of the para substituents and increase with decreasing basicity of the triarylphosphine. The rates of these reactions are also related to the reduction potentials (determined by cyclic voltammetry) of the azide complexes and increase with increasing ease of reduction. A mechanism is proposed for these reactions.

Spectrophotometric Determination of Iron(III) as Azide Complexes in Aqueous Tetrahydrofuran

Abstract A simple, sensitive and alternative method for the spectrophotometric determination of iron(III) has been established. The procedure is based on the formation of iron-azide complexes in 60% (v/v) tetrahydrofuran/ water medium. The high sensitivity obtained in this method is due to the use of an interesting absorption band not previously reported in the literature. In the recommended conditions, absorbances for the ferric complexes are measured at 400 nm where the molar absorptivity is 1.52 × 104 1 mol−1 cm−1. The organic solvent used increases the sensitivity and the stability of the measurements. The precision is shown by the average deviation of about 0.3%. This system obeys Beer's law and is suitable for iron(III) determination in the concentration range from 0.6 to 3.2 mg 1−1 (ppm). The best experimental conditions were determined studying the different factors involved. The influence of various diverse ions was also studied.

Determination of microgram amounts of azide by gas chromatography

Azide ions (5–120 μg) are oxidized by cerium(IV) in an acidic medium. The nitrogen evolved is displaced by helium carrier and quantified by gas chromatography with a thermal conductivity detector. The relative standard deviation is <5%. Many sulphur anions, cyanide and some metal ions which form azide complexes do not interfere.

Proton and nitrogen-15 NMR spectroscopic studies of hydrogen ion-dependent pseudo-halide ion binding to chloroperoxidase.

The proton nuclear magnetic resonance spectra of several chloroperoxidase-inhibitor complexes have been investigated. Titrations of chloroperoxidase with azide, thiocyanate, cyanate, or nitrite ions indicate that only the chloroperoxidase-thiocyanate complex exhibits slow ligand exchange on the 360-MHz NMR time scale. The temperature dependence of the proton NMR spectra of the complexes suggests that, although the complexes are predominantly low-spin ferric heme iron, a spin equilibrium is present presumably between S = 1/2 and S = 5/2 states. The pH dependence of the proton NMR spectra of the psuedo-halide-chloroperoxidase complexes was examined at 360 and 90 MHz. Chloroperoxidase complexes with azide and cyanate show similar behavior; 360-MHz proton spectra are readily observed at low pH (less than 5.0) but not at high pH. At high pH, the ligand exchange rate falls in an intermediate time range. When the complexes are examined at 90 MHz, however, spectra consisting of averaged signals are observed. The chloroperoxidase-thiocyanate complex does not form at high pH values; the proton NMR spectrum observed is that of native chloroperoxidase. The pKa for the chloroperoxidase-thiocyanate heme-linked ionizable amino acid residue falls between 4.2 and 5.0. Only an averaged azide signal was observed in the nitrogen-15 NMR spectra for solutions that contained the azide complex of chloroperoxidase, horseradish peroxidase, and myoglobin.

ChemInform Abstract: Potentiometric Study of Azide Complexes of Iron(III) in Aqueous Medium.

AbstractThe stepwise formation of Fe(III) ‐ azide complexes has been investigated a: 25 "C in 3 M NaClO.

Potentiometric study of azide complexes of iron(III) in aqueous medium

The stepwise formation of iron(III)-azide complexes in aqueous sodium perchlorate medium has been investigated at 25.0±0.1°C and ionic strength 3.0 M. By the indirect potentiometric method employed, using a glass electrode and under the conditions studied, five strong mononuclear complexes were detected. No evidence of polynuclear species was found. The final analysis of the data, by solution of simultaneous equations (matrix method), leads to the following overall formation constants: β 1 = 2.23 × 10 5 M −1, β 2 = 1.55 × 10 8 M −2, β 3 = 1.36 × 10 10 M −3, β 4 = 3.18 × 10 11 M −4 and β 5 = 3.11 × 10 12 M −5. The graphical method of Leden, with some refinements, was also utilized initially.

1H NMR spectroscopy of cytochrome cd1 derivatives

Proton nuclear magnetic resonance spectra are reported for cytochrome cd 1 from Pseudomonas aeruginosa (ATCC 19429) in several forms including complexes of the ferricytochrome with cyanide, azide, and fluoride, a quasi-apo form in which the noncovalently associated heme d 1 has been removed but the covalently bound heme c is retained, and the reduced state of both native and the quasi-apo forms. Comparisons are made to the previously reported spectrum of ferricytochrome cd 1. The following points are made. (i) The spectra of the azide and fluoride complexes and the ferric quasi-apo form show perturbation of resonances assignable to the site of heme d 1, and leave relatively unperturbed resonances assignable to the site of heme c. The heme d 1 associated resonances are at 46.0, 35.4, 23.3, 17.5, −2.9, and −16 ppm, and the heme c associated resonances are at 42.0, 33.7, 15.0, 13.9, −7.5, −14, and −33 ppm in native ferricytochrome cd 1. (ii) The similarity of the hyperfine resonances of the ferric quasi-apo form to the heme c resonances of intact ferricytochrome cd 1 is evidence that removal of heme d 1 leaves the heme c binding site relatively unaltered. (iii) Linewidths and relaxation times suggest that the relaxation times of the unpaired electron spins of the ferric hemes c and d 1 are on the same order of magnitude. (iv) Although it is paramagnetic, ferrocytochrome cd 1 does not demonstrate an experimentally detectable hyperfine shifted spectrum under present conditions. Possible reasons for this are discussed. (v) The presence of a narrow resonance at −2.8 ppm in both ferrocytochrome cd 1 and the reduced state of the quasi-apo form suggests that methionine may be a ligand to heme c.

Spin-label study of conformational changes induced by pH and ligands in leghemoglobin from yellow lupine root nodules.

Conformational changes induced by ligands and pH in lupine ferrileghemoglobin selectively modified at Tyr-E16 by the imidazolide spin label has been studied by the method of electron spin resonance in the pH range 6-13. It is shown that in the alkaline pH region the bound spin label registers a local conformational transition which precedes the alkaline denaturation of the protein. In aquamet, cyanide and nicotinate complexes of ferrileghemoglobin this transition occurs with pK 10.5, in acetate and azide complexes with pK 11. In all these ligand derivatives the transition is induced by alteration in the ionization state of one group (delta nH+ approximately equal to 1), most probably, the epsilon-amino group of Lys-GH3. The latter is linked with the Glu-A14 residue and this bond is essential for maintaining the native conformation of leghemoglobin. The ligand-induced conformational changes in the vicinity of the label are small and consist, most likely, in some alteration of the mutual arrangement of the AE and GH helical complexes. No correlation has been revealed between the spin state of the heme iron and the conformation of leghemoglobin in the region under study.

Thermochromism of the azide complexes of ferric hemoproteins

The thermochromism of ferric azide hemoglobin and myoglobin was examined in a 0–35°C range and the visible spectra of the two spin isomers were separated from the analysis of the thermochromism. (1) The separated low-spin spectra exhibit intensified 540 and 570 nm peaks and a red-shifted Soret band. The high-spin spectra show characteristic 4-banded visible absorption, which is significantly different from the spectrum of 5-coordinate azide hemin, suggesting that the heme iron of azide hemoprotein remains 6-coordinate during the spin-state transition. (2) A larger thermochromism was observed in carp azide methemoglobin bound with inositol hexaphosphate, in contrast with the results reported on human hemoglobin. The difference between the human and carp hemoglobins was consistently explained in terms of an equilibrium coexistence of the R and T conformers in human azide methemoglobin bound with inositol hexaphosphate.

Complexes of cytochrome caa3 from the thermophilic bacterium PS3 formed with ligands and during catalytic activity.

PS3 (thermophilic bacterium) cytochrome caa3 reacts slowly with cyanide which forms a low-spin complex with the CuBa3 centre. Partial reduction under catalytic conditions increases the rate of cyanide binding, and the reaction constant is rather similar to that of the mammalian enzyme, but the partially reduced complex dissociates more rapidly than does the corresponding eukaryotic complex. A simple biphasic reaction can account for the results obtained. The azide complex of partially reduced PS3 cytochrome caa3 shows an alpha-peak blue shift similar to that of the mammalian enzyme. PS3 cytochrome caa3 forms an oxyferri ("oxygenated") species like the mammalian enzyme, but does not undergo high- to low-spin changes during the aerobic steady state with ascorbate and N,N,N',N'-tetramethyl-p-phenylenediamine as substrates. Interactions seen in cytochrome oxidase-ligand reactions and its spin-state changes are therefore intrinsic to the enzyme's large catalytic subunits and do not require the presence of the small nuclear-encoded subunits found in eukaryotic systems.

Azide mit zwei verschiedenen Metallatomen

Crystal structures of azides containing two different metal atoms are discussed. Elements with higher electronegativity have the greatest influence on the structures. They form finite or infinite metal azide complexes, the degree of covalent binding betweenM(II) and azide groups raises with increasing ionisation potential of the metal, the stability of complex azides is not influenced seriously thereof. The arrangement ofM(I)-ions is influenced to a large extent by the stereochemistry. The azide groups are essentially asymmetrical with N-N-distances between 1.10 and 1.28 A, the mean distance is 1.18 (3) A. Azide groups are slightly bent [N-N-N=177 (5)°] and are always coordinated to several metal atoms. Frequently the central nitrogen atoms of azide groups are located closer toM(I)-ions than terminal nitrogens. Some complex azides contain water which is situated beside the azide groups within the coordination polyhedra.

Spectrophotometric Determination of Copper(II) as the Azide Complexes, in the Presence of Iron(III) Cations

Abstract A method for the spectrophotometrio determination of copper(II), in the presence of iron(III) cations (excess), was stablished. The masking of iron is made with sodium fluoride salt in 50 % (v/v) water/acetone medium. In the recommended conditions, absorbances for cupric complexes are measured at 435 nm where molar absorptivity is 6.00 × 103 1 mol−1 cm−1. The stable ayetern obeys Beer's law and is suitable for the copper determination in concentration range from 2.0 to 9.0 mg 1−l. The iron(III) ion interference (until ca. 600 mg 1−l) can be completely suppressed. The influence of diverse ions and several others factore were studied. The results show that copper(II) can be accurately determined by azide apectrophotometric method, if the samples were suitablely treated by the recommended procedure.

Azides of heavier main group elements: The reluctance of phosphine azides to undergo a curtius-type rearrangement

Under thermolytic or photolytic conditions, phosphine azide complexes Ph 2PN 3M(CO) 5 (M = W, Cr) gave rise to the corresponding isocyanato complexes and not to the expected iminophosphane complexes.

Metal site cooperativity within cytochrome oxidase.

Low temperature (9-15 K) EPR of isolated bovine heart cytochrome oxidase titrated potentiometrically in the presence of azide reveals the formation of two distinct species of low-spin cytochrome a3(III)-azide which differ in redox properties and g values. Both species are formed with characteristic midpoint potentials during the course of oxidative titration and disappear at higher potentials. The signal appearing at lower potential has principal g values 2.88, 2.19, and 1.64; that appearing at higher potential has g values 2.77, 2.18, and 1.74. A good fit to the experimental data (per cent of cytochrome present in a given paramagnetic state versus oxidation potential) was obtained with a model whereby the gz = 2.88 species arises from cytochrome a3(III)-azide with cytochrome a reduced, which is converted to the gz = 2.77 species upon oxidation of cytochrome a. Potentiometric titration of cytochrome oxidase in the presence of cyanide produces two low-spin heme EPR signals attributable to cytochrome a3(III)-cyanide which are incompletely resolved, but are distinguishable nonetheless. The low-potential signal has peak amplitude at gz = 3.63 and a long high-field tail; this resonance has been seen by other workers in the partially reduced enzyme (DerVartanian, D. V., Lee, I. Y., Slater, E. C., and van Gelder, B. F. (1974) Biochim. Biophys. Acta 347, 321-327). The high-potential signal is much more symmetric about its peak amplitude, which is at approximately 10 G higher field with gz = 3.61. As with the azide complex, the titration behavior in the presence of 2 mM KCN is adequately simulated by assuming that the appearance of the two species is a function of the oxidation state of cytochrome a. Like the a3-azide signals, the a3-cyanide signals disappear upon further oxidation with some characteristic midpoint potential. If the disappearance of the a3-ligand signals with increasing potential is assumed to be the result of antiferromagnetic (or ferromagnetic) coupling of a3(III) (S = 1/2) to CuB(II) (S = 1/2), then cooperativity between cytochrome a and CuB is implied. The data are consistent with the hypothesis that oxidation of cytochrome a raises the midpoint potential of CuB by 55 +/- 10 mV.

Iron(III) complexes of sodium 2-pyridinecarboxaldehyde dithiocarbazonate, a possible chelating agent for the treatment of iron overload

The 1:2 complexes of iron(III) with sodium 2-pyridinecarboxaldehyde dithiocarbazonate of the formula Fe(PD) 2X (where PD is 2-pyridinecarboxaldehyde dithiocarbazonate anion, and X − = Cl −, NO 3 −, N 3 − and NCS −) have been prepared by interaction of iron(III) ions with NaPD (1:2), in the presence of an appropriate anion. They show molar conductance values of 33.0 to 55.2 mhos cm 2 mol −1 in DMF solution. These data indicate that the anions of FE(PD) 2X are associated in the first coordination sphere of Fe(III) to a different extent in DMF solution. The ultraviolet-visible absorption spectral data of FE(PD) 2X in DMF solution also support the different extent of coordination of anions to Fe(III). The Fe(PD) 2X complexes in the solid state have low spin iron(III) with μ eff values varying from 1.95 to 2.35 BM. The EPR data of the iron(III) complexes in frozen DMF also support their low-spin nature. These data also show that all complexes have rhombic symmetry except the azide complexes which have axial symmetry. The EPR and infrared spectral data of iron(III) complexes also suggest that the anions of these complexes are associated in the first coordination sphere of iron(III) in the solid state. The association of anions in the Fe(PD) 2X complexes may suggest their geometry to be capped octahedron or pentagonal bipyramid. It has been suggested that one of the most important factors for efficiently removing iron from iron loaded mice is the formation of kinetically inert low-spin iron(III) complexes between iron(III) ions and tridentate chelating agents in the ratio of 1:2.

Azide interaction with 4f and 5f ions in aqueous solutions. 1. Trivalent ions

Solvent extraction and UV, Raman, and NMR spectroscopic studies were carried out on trivalent actinide and lanthanide aqueous azido complexes. Unlike trivalent d transition ions (..beta../sub 11/ approx. = 10/sup 5/) 5f and 4f aqueous azido complexes are weak (..beta../sub 11/ approx. = 2.5 for neodymium azide complexes (Nd(N/sub 3/)/sup 2 +/)), but the trivalent actinides exhibit formation constants 1 order of magnitude higher than the lanthanides. All the spectroscopic methods indicate that we are dealing with inner-sphere complexes and actinide-lanthanide differences must be attributed to higher covalent contributions in the 5f azides. /sup 15/N NMR combined with /sup 1/H NMR served to investigate the azide binding properties. With the trivalent 5f and 4f ions the binding occurs by one of the terminal nitrogen atoms. The M-N bond distance is close to 2.75 A. The lanthanide(III)-(linear azide) moieties are bent with a bond angle close to 135/sup 0/, unlike the homologous linear thiocyanate complexes. 7 figures, 4 tables.

Simultaneous spectrophotometric determination of copper(II) and iron(III) as the azide complexes

The simultaneous spetrophotometric determination of copper and iron is based on the yellow and red azide complexes formed in 50% (v/v) water/acetone medium. Absorbances are measured at 345 and 435 nm where molar absorptivities for the iron(III) complex are 8.77 × 10 3 and 8.49 × 10 3 l mol −1 cm −1, rspectively, and absorptivities for the copper(II) complexes are 1.47 × 10 3 and 5.69 × 10 3 l mol −1 cm −1, respectively. The systems obey Beer's law and concentrations as low as 2.0 and 1.2 mg l −1 for copper and iron, respectively, can be quantified in a 1.00-cm cell. Determination of iron(III) in 23 mixtures (1.1–5.4 mg l −1) yielded a relative standard deviation of 0.86%; similar data for copper(II) (2.0–8.9 mg l −1) yielded a relative standard deviation of 1.6%.

Model complexes for the active sites of reduced and oxidized sites of hemocyanin and tyrosinase. Structures of binuclear Cu(I) and Cu(II) complexes and characterization of a model copper monooxygenase reaction

It has been established that the dioxygen carrying copper containing protein hemocyanin and the copper monooxygenase tyrosinase both contain electronically coupled binuclear copper centers. In the deoxy protein, copper centers are thought to be two or three coordinate, while in their oxidized form, tetragonally coordinated Cu(II) ions are bridged by an endogenous protein group which is thought to be phenolate from tyrosine. Coordinated dioxygen (as peroxide) is also known to bridge the two coppers. The binding of other exogenous ligands (azide, chloride, etc.) to the binuclear copper centers has been used to probe the active site structures and some of these may also bridge the Cu(II) ions in methemocyanin or tyrosinase derivatives [1]. Extensive coordination model studies are being carried out [2–4] in order to 1) determine the nature of the ligand donors and coordination geometries of these active site copper centers 2) mimick and understand the coordination chemistry of relevant Cu(I) systems; 3) mimick the Cu(II) binuclear centers in order to relate the spectroscopic and chemical properties of these models and their derivatives (with appropriate probes) to those of the proteins and 4) understand those factors necessary and the mechanism(s) for reversible dioxygen binding and ‘activation’ of dioxygen ( i.e. specific hydroxylation of substrates by the copper monooxygenases). As part of our own extensive investigations into the chemistry of binuclear Cu(I) and Cu(II) moieties, we have employed the binucleating ligand m-XYLpy, I (py = 2-pyridyl), where two tridentate donor groups are separated by a meta-xylyl bridge [5]. A novel binuclear cuprous complex, II, ( See next column) containing well separated trigonally coordinated Cu(I) ions is isolated by the reaction of I with two equivalents of Cu(CH 3CN) 4PF 6 [6]. The coordination geometry and the type of ligands in II makes it a good model complex for the deoxy-sites in the proteins. In addition, II reacts with O 2 in a manner analogous to the copper monooxygenase tyrosinase. Specific hydroxylation of the xylyl ligand occurs producing the phenoxy- and hydroxy- (R = H) doubly bridged binuclear Cu(II) complex III. Manometric O 2 uptake experiments with II and mass spectrometric analyses of the oxidation product III utilizing isotopically labeled dioxygen demonstrate that both atoms of oxygen in III are derived from 18O 2[6, 7]: ▪ The structures of compounds II and III have been proven by X-ray crystallographic studies [5–7]. Compounds III contain tetragonally coordinated Cu(II) ions which are phenolate bridged (Cu⋯Cu = ∼3.1 Å), making them excellent structural models for the proposed active sites in hemocyanin and tyrosinase. Reaction of III (R = H) with HX (X = N 3 − , Cl − results in the replacement of the bridging ligand (-OH) by X with the formation of complexes IV. These have also been structurally characterized and they exhibit coordination geometries very similar to III. In the case of the azide complex (X = N 3 − ), a novel μ-1,1-azido type of bridging is observed [7]. This mode of coordination has not been previously proposed to take place in the protein copper centers. Its possible occurrence should be considered in studies where azide ligand binding is used as a spectral probe of the protein copper centers.