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Microbial cells are well known to be capable of remaining viable when desiccated, and a variety of beneficial microorganisms can thus be preserved for storage. For the ubiquitous widely studied soil bacterium Azospirillum brasilense (wild-type strain Sp7), which has a significant agrobiotechnological potential owing to its plant-growth-promoting capabilities perspective for its use in biofertilisers, Fourier transform infrared (FTIR) spectroscopy (in the diffuse reflectance mode, DRIFT) was used to control the state of biomass, together with 57Fe transmission Mossbauer spectroscopy to monitor intracellular iron speciation in live rapidly frozen cell suspension and in the lyophilised biomass (both measured at T = 80 K). It has been shown for the first time that a relatively large part of ferrous iron in live cells (22% of the whole cellular iron pool, represented by two high-spin Fe(II) forms, in the 18-h culture grown on 57Fe(III) complex with nitrilotriacetic acid as the sole source of iron) gets largely oxidised upon lyophilisation. The remaining part of iron(II) in the resulting dry biomass was found to be ca. 3% only. The major part of ferric iron in the dry biomass was shown to be comprised of ferritin-like ferric species (giving a typical magnetically split sextet at T = 5 K), while the iron(III) formed from cellular iron(II) by oxidation in air in the course of drying remained in a paramagnetic state even at T = 5 K. The possibility of intracellular iron(II) oxidation to iron(III) upon desiccation may be a specific natural strategy to avoid cell damage caused by Fenton-type reactions in dormant (frozen, dried) cells. The results obtained may have important implications related to iron speciation and redox transformations in dried bacterial preparations intended for long-term storage.

In this review, the unique possibilities are considered of the 57Fe transmission (TMS) and 57Co emission (EMS) variants of Mossbauer (nuclear γ-resonance) spectroscopy as nondestructive techniques with minimal sample preparation/treatment and a significant analytical potential, with a focus on the analysis of cation-binding sites in metalloproteins. The techniques are shown to provide unique structural and quantitative information on the coordination microenvironment, the chemical state and transformations of the Mossbauer nuclides in sophisticated metal-containing proteins, including those within complicated supramolecular structures, and in microbial cells or tissues. Recent representative examples of analyses of Fe-containing proteins by 57Fe TMS are briefly discussed, along with the newly emerging data on using 57Co EMS for probing the structural organisation of 57Co-doped cation-binding sites in sophisticated biocomplexes including metalloenzymes. Finally, some rare or exotic applications of Mossbauer spectroscopy (including the synchrotron-based methodology) in protein-related studies are outlined.

In the emission (57Co) variant of Mossbauer spectroscopy (EMS), the 57Co radionuclide (with a half-life of 9months) is used that undergoes a nuclear decay 57Co→57Fe via electron capture followed by the emission of a γ-quantum, the energy of which is modified by the chemical state and the close coordination environment of the parent 57Co atom. While EMS has been used largely in materials science and nuclear chemistry, its high sensitivity can also be of great advantage in revealing fine structural features and for speciation analysis of biological complexes, whenever the 57Co2+ cation can be used directly as the coordinating metal or as a substitute for native cobalt or other metal ions. As such EMS applications are yet rare, in order to reliably interpret emission spectra of sophisticated 57Co2+-doped biosystems, model EMS studies of simple cobalt biocomplexes are necessary. In this work, EMS spectroscopic data are analysed and discussed for 57Co2+ complexes with a range of small biomolecules of different structures, including 4-n-hexylresorcinol, homoserine lactone and a few amino acids (spectra measured in rapidly frozen dilute aqueous solutions or in the dried state at T=80K). The EMS data obtained are discussed with regard to the available literature data related to the coordination modes of the biocomplexes under study.

For the ubiquitous diazotrophic rhizobacterium Azospirillum brasilense, which has been attracting the attention of researchers worldwide for the last 35 years owing to its significant agrobiotechnological and phytostimulating potential, the data on iron acquisition and its chemical speciation in cells are scarce. In this work, for the first time for azospirilla, low-temperature (at 80 K, 5 K, as well as at 2 K without and with an external magnetic field of 5 T) transmission Mössbauer spectroscopic studies were performed for lyophilised biomass of A. brasilense (wild-type strain Sp7 grown with (57)Fe(III) nitrilotriacetate complex as the sole source of iron) to enable quantitative chemical speciation analysis of the intracellular iron. In the Mössbauer spectrum at 80 K, a broadened quadrupole doublet of high-spin iron(III) was observed with a few percent of a high-spin iron(II) contribution. In the spectrum measured at 5 K, a dominant magnetically split component appeared with the parameters typical of ferritin species from other bacteria, together with a quadrupole doublet of a superparamagnetic iron(III) component and a similarly small contribution from the high-spin iron(II) component. The Mössbauer spectra recorded at 2 K (with or without a 5 T external field) confirmed the assignment of ferritin species. About 20% of total Fe in the dry cells of A. brasilense strain Sp7 were present in iron(III) forms superparamagnetic at both 5 and 2 K, i.e. either different from ferritin cores or as ferritin components with very small particle sizes.

The transmission (57Fe) and emission (57Co) variants of Mossbauer spectroscopy are used to monitor metabolic transformations of 57FeIII/57FeII and 57CoII in cells of the bacterium Azospirillum brasilense Sp7. (The spectra are measured at 80 K.) For 57CoII, metabolic changes in the living cells are observed after 1 h. Two components of high-spin 57FeII are observed for the bacteria grown with 57FeIII as a source of iron; the parameters for 57FeIII correspond to ferritin-like components.

The results of a comparative study of two samples of the rhizobacterium Azospirillum brasilense (strain Sp245) prepared in different conditions and of human liver ferritin using Mossbauer spectroscopy with a high velocity resolution demonstrated the presence of ferritin-like iron (i.e. iron similar to that found in ferritin-like proteins) in the bacterium. Mossbauer spectra of these samples were fitted in two ways: as a rough approximation using a one quadrupole doublet fit (the homogeneous iron core model) and using a superposition of quadrupole doublets (the heterogeneous iron core model). Both results demonstrated differences in the Mossbauer parameters for mammalian ferritin and for bacterial ferritin-like iron. Moreover, some differences in the Mossbauer parameters were observed between the two samples of A. brasilense Sp245 related to the differences in their preparation conditions.

Iron(III)-containing aqueous solutions of 5-methylresorcinol (5-MR), 5-n-propylresorcinol (5-n-PR) and 4-n-hexylresorcinol (4-n-HR) at pH ~ 3 were studied by means of 57Fe transmission Mossbauer spectroscopy. Kinetic considerations were applied to the redox reactions. Density Functional Theory (DFT) calculations were performed for the alkylresorcinol (AR) molecules and their non-alkylated analogue (resorcinol). Mossbauer spectra consisted of quadrupole doublets assigned to high-spin Fe(III) and Fe(II) species. From changes in their relative spectral areas, a gradual reduction of Fe(III) by all the ARs studied was observed. However, significant differences were found for the reduction rates among the ARs. The following series of the reduction rates was established by means of Mossbauer spectroscopy: 4-n-HR ≫ 5-MR > 5-n-PR, supplemented by rate constants calculated using a kinetic model. DFT calculations resulted in the following series: 4-n-HR ≫ 5-n-PR > 5-MR ≫ resorcinol (the latter is not oxidised under the conditions applied). The reversed order of the experimentally observed 5-MR and 5-n-PR oxidation rates may be explained in terms of their different kinetic parameters related to their structure.

Preliminary 57 Fe transmission Mossbauer spectroscopic data were ob- tained for the first time for live cells of the plant-growth-promoting rhizobacterium Azospirillum brasilense (wild-type strain Sp245) grown aerobically with 57 Fe III - nitrilotriacetate (NTA) complex as a sole source of iron. The results obtained have shown that live cells actively reduce part of the assimilated iron(III) to iron(II), the latter amounting up to 33 % of total cellular iron after 18 h of growth, and 48 % after additional 3 days of storage of the dense wet cell suspension in nutrient-free saline solution in air at room temperature (measured at 80 K). The cellular iron(II) was found to be represented by two quadrupole doublets of different high-spin forms, while the parameters of the cellular iron(III) were close to those typical for bacterioferritins.

Mossbauer spectroscopy is used for a comparative study of the rate of iron(III) reduction by 4-n-hexylresorcinol (4-n-HR, a chemical analog of microbial autoregulators excreted by cells into the environment that allow intercellular communication) in aqueous media in the pH range of 1.5–4.5 simulating acidic soil conditions. The concomitant process of 4-n-HR oxidation is monitored using UV spectrophotometry.

Thermoplastic linear ionomer based on sulphonated poly(ether-urethane)—styrene-acrylate copolymer, doped with natural Fe2 + , was studied by Mossbauer spectroscopy at T = 78 and 290 K to monitor the chemical state of Fe species. The Fe2 + added to aqueous suspension of the system was only partly oxidised in the course of polymer film preparation and drying in air. The oxidised part comprised a magnetic phase (~19 % of total Fe both at T = 78 and 298 K) and a quadrupole doublet (~40 %), while FeII (over 40 %) stabilised in two types of microenvironments.

Emission (57Co) Mossbauer spectra of the aspartic acid—57CoCl2 system were measured at T = 80 K in frozen aqueous solution and in the form of a dried residue of this solution. The Mossbauer spectra, besides a weak contribution from after-effects, showed two Fe2 + /Co2 + components which were ascribed to octahedrally and tetrahedrally coordinated 57CoII microenvironments in the Asp–cobalt(II) complex. This dual coordination mode may be due to the involvement of the second terminal carboxylic group of aspartic acid in the coordination sphere of Co.

Mossbauer spectroscopy was used to study iron(III) reduction by alkylresorcinols (ARs; i.e., chemical analogs of microbial autoregulators excreted by cells into the environment and involved in intercellular communication) in aqueous solutions simulating acidic soil conditions. The concomitant oxidation process for an AR (5-methylresorcinol) was monitored using UV spectrophotometry.

Thermoplastic linear poly(ether-urethane) sulphonated ionomer (PEUSI), doped with natural Fe2 + , was studied by Mossbauer spectroscopy at T = 78 and 290 K in order to monitor the chemical state of Fe species. The Fe2 + added was found to get only partly oxidised in the course of preparation of a dry polymer film from aqueous suspension. The oxidised part gave a ferric quadrupole doublet (over half of total Fe added) and, at T = 78 K only, a small portion of magnetically split sextet (ca. 3%), whereas FeII (total over 40%) stabilised in two different forms (represented by quadrupole doublets) corresponding to two types of iron(II) microenvironments, where FeII could be coordinated involving –CONH– and –NHNH– moieties and sulphonate groups in PEUSI. The results obtained show that Fe2 + -doping of PEUSI-based materials may be used for probing their structural characteristics by Mossbauer spectroscopy.

The influence of water on the interaction between iron(III) and indole-3-acetic acid (IAA) was studied in different organic solutions using rapid-scan stopped-flow spectrophotometry and rapid-freeze/quench Mossbauer spectroscopy. Measurements were also performed in ethanol–water and acetone–water mixtures. The results showed that the interaction between FeIII and IAA resulted in dimeric FeIII–IAA complex within 1 s, followed by a slow second step to give Fe2+ and IAA(oxidized). No such products were formed in the absence of water. The visible and Mossbauer spectra reflect the nature of the organic solvent and that of the anion of iron(III) salts.

The chemical reactions between iron(III) and indole-3-acetic (IAA), -propionic (IPA), and -butyric (IBA) acids were studied in acidic aqueous solutions. The motivation of this work was that IAA is one of the most powerful natural plant-growth-regulating substances (phytohormones of the auxin series). Mossbauer spectra of the frozen aqueous solutions of iron(III) with indole-3-alkanoic acids as ligands (L), showed parallel reactions between Fe3+ and the ligands. Partly, it resulted in a complex formation which precipitated in aqueous solution and partly, in a redox process with iron(II) and the oxidised indole-3-alkanoic acids as products. The Mossbauer parameters of the Fe2+ species suggested a hexaaquo coordination environment. The chemical composition and coordination structure of the precipitated complexes were investigated using elemental analysis, Mossbauer spectroscopy, Fourier transform infrared (FTIR) and Raman spectroscopic techniques. The complexes were soluble in some organic solvents. So, Mossbauer, FTIR and solution X-ray diffraction measurements were carried out on the solution of complexes in acetone, hexadeutero acetone and methanol, respectively. The data obtained supported the existence of the μ-dihydroxo-bridging structure of the dimer: [L2Fe FeL2] (where L is indole-3-propionate, -acetate or -butyrate).

This paper describes the Mossbauer investigations of iron(III) salts in aqueous solutions in the presence of indole-3-alkanoic acid ligands. The measurements showed two parallel reactions between the ligands and ferric ions: a complex formation and a redox process. The oxidation process takes place in the ligands, and a part of Fe3+is reduced to Fe2+.

Emission Mossbauer spectroscopy with the 57Co isotope was used to study very dilute rapidly frozen aqueous solutions of cobalt(II) complexes with low-molecular-weight biomolecules (aromatic amino acids — anthranilic acid and L-tryptophan) and within a sophisticated biopolymer, bacterial glutamine synthetase, a key enzyme of nitrogen metabolism. The appearance of after-effects of the 57Co→57Fe nuclear transformation as well as the coordination properties of the cation and the ligands in the complexes are discussed on the basis of their Mossbauer parameters.

Cobalt is an essential trace element with a broad range of physiological and biochemical functions. However, biochemical speciation of cobalt and structural investigations of cobalt-containing complexes with biomacromolecules are challenging, as the participation of cobalt in physiological processes is limited by its very low concentrations. Emission Mossbauer spectroscopy (EMS), with the radioactive 57 Co isotope as the most widely used nuclide, is several orders of magnitude more sensitive than its 57 Fe absorption variant which has had a rich history of applications in bioscience. Nevertheless, owing to specific difficulties related to the necessity of using radioactive 57 Co in samples under study, applications of EMS in biological fields have so far been sparse. In this communication, the EMS applicability to studying biological objects as well as some specific aspects of the EMS methodology are considered in order to draw attention to the unique structural information which can be obtained non-destructively in situ. Chemical consequences (after-effects) of the nuclear transition ( 57 Co→ 57 Fe), which provide additional information on the electron acceptor properties of the proximate chemical microenvironment of the metal ions, are also considered. The data presented demonstrate that EMS is a sensitive tool for monitoring the chemical state and coordination of cobalt species in biological matter and in biomacromolecular complexes (metalloenzymes), providing valuable structural information at the atomic level.

CD spectroscopic study of the secondary structure of partly adenylylated glutamine synthetase (GS) of the bacterium Azospirillum brasilense showed both the native and cation-free (EDTA-treated) enzyme to be highly structured (58 and 49% as α-helices, 10 and 20% as β-structure, respectively). Mg2+, Mn2+, or Co2+, when added to the native GS, had little effect on its CD spectrum, whereas their effects on the cation-free GS were more pronounced. Emission (57Co) Mossbauer spectroscopic (EMS) study of 57Co2+-doped cation-free GS in frozen solution and in the dried state gave similar spectra and Mossbauer parameters for the corresponding spectral components, reflecting the ability of the Co2+–enzyme complex to retain its properties upon drying. The EMS data show that (a) A. brasilense GS has 2 cation-binding sites per active center and (b) one site has a higher affinity to Co2+ than the other, in line with the data on other bacterial GSs. © 2004 Wiley Periodicals, Inc. Biopolymers, 2004

Mossbauer spectroscopic studies were carried out in acidic (pH 2.3) 57FeIII nitrate containing aqueous solutions of indole-3-butyric acid (IBA), rapidly frozen in liquid nitrogen at various periods of time after mixing the reagents. The data obtained show that in solution in the presence of IBA, iron(III) forms a complex with a dimeric structure characterised by a quadrupole doublet, whereas without IBA under similar conditions iron(III) exhibits a broad spectral feature due to a slow paramagnetic spin relaxation which, at liquid nitrogen temperature, results in a large anomalous line broadening (or, at T = 4.2 K, in a hyperfine magnetic splitting). The spectra of 57FeIII+IBA solutions, kept at ambient temperature under aerobic conditions for increasing periods of time before freezing, contained a gradually increasing contribution of a component with a higher quadrupole splitting. The Mossbauer parameters for that component are typical for iron(II) aquo complexes, thus showing that under these conditions gradual reduction of iron(III) occurs, so that the majority (85%) of dissolved iron(III) is reduced within 2 days. The Mossbauer parameters for the iron(III)-IBA complex in aqueous solution and in the solid state (separated from the solution by filtration) were found to be similar, which may indicate that the dissolved and solid complexes have the same composition and/or iron(III) coordination environment. For the solid complex, the data of elemental analysis suggest the following composition of the dimer: [L2Fe-(OH)2-FeL2] (where L is indole-3-butyrate). This structure is also in agreement with the data of infrared spectroscopic study of the complex reported earlier, with the side-chain carboxylic group in indole-3-butyrate as a bidentate ligand. The Mossbauer parameters for the solid 57FeIII-IBA complex at T = 80 K and its acetone solution rapidly frozen in liquid nitrogen were virtually identical, which indicates that the complex retains its structure upon dissolution in acetone.

FeIII was found to be gradually reduced by indole-3-acetic acid (IAA) in slightly acidic nitrate solution with further re-oxidation of the resulting FeII upon drying in air, which is reasoned to be of ecological significance considering IAA excretion by many soil micro-organisms. FeIII–IAA complex was isolated; its composition, some physicochemical properties and structural aspects were studied using Mossbauer, FTIR spectroscopic and other techniques. Possible influence of FeIII coordination on the redox properties of the ligand is also considered from the viewpoint of the nature of the bonding system which involves the conjugated π-electronic system of the pyrrole cycle along with the carboxylic O-donor atom.

Mossbauer, FTIR and XRD analyses showed that in aqueous medium in air in the presence of L-tryptophan (Trp) or indole-3-acetic acid (IAA) the ambient-temperature ageing of the precipitates formed from ferrous sulphate at pH∼7 gave composite phases with varying proportions of γ-FeOOH (a dominating crystalline phase), α-FeOOH (both fine-grained, showing superparamagnetic behaviour at 298 K, and relatively better crystallized) and amorphous ferric hydroxide. The experimental data suggested a competition for adsorption sites at the oxyhydroxide surface in the suspension during phase transformations, as well as the transformation of γ-FeOOH (and/or amorphous ferric hydroxide) to α-FeOOH via the dissolution-reprecipitation mechanism. The formation of certain ferric oxyhydroxide phases in the presence of Trp and IAA — released e.g., in the course of bacterial and plant metabolism — can contribute to the regulation of soil mineral composition.

The data on the coordination and structure of iron(II) and iron(III) anthranilates in the solid state and in aqueous medium are presented and discussed, as studied using Mossbauer (for solid products and frozen solutions) and FTIR spectroscopy (for solid samples). Whereas, in slightly acidic nitrate solutions under aerobic conditions ferric ions can still be gradually reduced by anthranilic (o-aminobenzoic) acid, which may have some ecological significance, in circumneutral media this process is retarded. Mossbauer parameters calculated for iron(II) and iron(III) anthranilates, as well as characteristic vibration modes of certain functional groups involved in coordination with iron cations are discussed. The FTIR data obtained for ferrous anthranilates, as compared to anthranilic acid, definitely exhibit the direct involvement of both the carboxylic and the amino groups of anthranilic acid in coordination with iron. q 1999 Elsevier Science B.V. All rights reserved.

Fourier transform infrared (FTIR) spectroscopic measurements, performed in the far-IR region (200–800 cm−1 on binary coprecipitated Ni(II)-Fe(III) hydroxides (20–60 at.% Fe ( Ni + Fe ) ), are presented and discussed. Heating the sample containing 60 at.% Fe to 600°C, monitored by differential scanning calorimetry (DSC) measurements, results in the formation of a nickel ferrite structure from its precursor present in binary hydroxides. The role of Fe-to-Ni ratio non-uniformities within the surface and bulk of the binary hydroxide crystallites, as revealed by Mossbauer measurements, Auger electron spectroscopy and depth profiling performed previously, is also discussed with regard to the formation of α-Fe2O3 and NiO impurities on thermal treatment.

The data of Mössbauer spectroscopic measurements in rapidly frozen 57FeIII nitrate solutions containing indole-3-acetic acid (IAA) are presented. The results obtained provide direct evidence that iron(III) is gradually reduced by IAA with the formation of soluble iron(II) complex. It has been found that further drying of the solution in air results in complete re-oxidation of iron(II) with the formation of iron(III) complex. The structure of the complexes obtained and their possible role in solubilization and transformation of iron species in soil in the presence of IAA secreted by soil microorganisms into the environment are also discussed.

The data of Mossbauer spectroscopic measurements in rapidly frozen acidified 57 Fe III nitrate solutions containing anthranilic acid are presented and discussed. The results obtained provide direct evidence that, along with the formation of an iron(III) anthranilate complex, gradual reduction of iron(III) by anthranilic acid occurs with the formation of iron(II) species even in acidic nitrate media.

In situ room-temperature transmission Mossbauer spectroscopic study of precipitates formed in 57 Fe(III)-containing highly alkaline electrolytes has been carried out after different periods of their ageing in the same solution. After 3.5 years of ageing, an intensive resonant absorption was detected in the form of a single line. The latter effect has not been observed earlier for solid iron(III) hydroxo derivatives which have commonly been expected to give either quadrupole or hyperfine magnetic splitting. The precipitate aged for 15 months was found to give an essentially lower Mossbauer effect. After complete hydrolysis in water both products gave comparably intensive quadrupole-split doublets with distinctly different parameters owing to differences in the structures of the initial phases. The absence of quadrupole splitting for the 3.5-year-aged product is interpreted in terms of the formation of the cubic symmetry of the ferric ion coordination polyhedron in sodium hydroxoferrate(III), slowly forming in the course of long-term storage in highly alkaline medium, while at earlier stages of ageing, the precipitate consists of a labile, possibly intermediate, phase ( eg ultrafine alpha-FeOOH particles) characterized by a low recoilless absorption probability. The results obtained are considered in relation with the processes occurring in alkaline storage batteries and electrolysers. Some effects leading to Mossbauer linewidth broadening, which may in principle take place in strongly alkaline electrolytes containing ferric derivatives with a symmetric hydroxo coordination, are also discussed.

Transmission Mossbauer and Fourier transform infrared spectroscopic data obtained for binary nickel(II)-iron(III) hydroxide systems in a broad range of compositions (3 to 60 at.% Fe (Ni+Fe) ) are compared. Structural transformations of the samples and their surface layers are considered, depending on their bulk gross compositions. The formation of a nickel ferrite precursor is shown to be promoted in the crystallites surface layers even at much lower gross Fe-to-Ni ratios than that in NiFe2O4 owing to surface enrichment with iron, as confirmed by Auger electron spectroscopic analyses.

A brief overview is given of recently obtained data on the surface structure and composition of various iron(III)-containing nickel hydroxide systems, as studied using transmission Mossbauer spectroscopy supplemented by atomic absorption, Auger electron and Fourier transform infrared spectroscopic data. The systems studied include coprecipitated binary Ni(II)−Fe(III) hydroxides in a wide range of compositions, ferric hydroxide precipitated on the surface of β-Ni(OH)2 crystallites, ferric hydroxo complexes adsorbed onto nickel hydroxides from alkaline electrolytes and the products of their hydrolysis.

The emission ((57)Co) variant of Mössbauer spectroscopy, rarely used in biology-related studies, was applied to study binding and possible transformations of (57)Co(II) traces in live and dead (hydrothermally treated) cells of the rhizobacterium Azospirillum brasilense (strain Sp7) at T=80 K in frozen aqueous suspensions and as their dried residues. The Mössbauer parameters calculated from the spectra were compared with the similarly obtained data reported earlier for another A. brasilense strain, Sp245 (which differs from strain Sp7 by the ecological niche occupied in the rhizosphere and was found earlier to exhibit different metabolic responses under similar environmental conditions). Similarly to strain Sp245, live cells of strain Sp7, rapidly frozen 2 min and 1 h after their contact with (57)Co(2+) (measured in frozen suspensions), showed marked differences in their Mössbauer parameters, reflecting metabolic transformations of (57)Co(2+) occurring within an hour. However, the parameters for strains Sp7 (this work) and Sp245 (reported earlier), obtained under similar conditions, were found to significantly differ, implying dissimilarity in their metabolic response to Co(2+). This is in line with their different metabolic responses to several heavy metals, including Co(2+), detected earlier using Fourier transform infrared spectroscopy.

An in situ transmission Mossbauer spectrum in the form of an intensive symmetric singlet has been obtained for a precipitate, formed in 57 Fe(III)-containing highly alkaline NaOH electrolyte, after its long-term ageing under the latter solution, which has not been reported earlier for solid ferric hydroxo derivatives. Hydrolysis of the sample in water leads to the formation of amorphous ferric hydroxide giving a quadrupole-split spectrum with a significantly lowered reasonant absorption intensity. The results obtained are interpreted in terms of the formation of sodium hydroxoferrate(III) featured by the cubin symmetery of the iron(III) coordination polyhedron, which is stable in concentrated alkali, giving an amorphous product upon hydrolysis.

Surface analysis of coprecipitated binary Ni(II)-Fe(III) hydroxide solid solutions using Auger electron spectroscopy and depth profiling has revealed redistribution of the components (nickel and iron) within the surface layers showing significantly increased iron-to-nickel ratios, as compared to the corresponding gross averaged values in the bulk. The results are compared with transmission Mossbauer spectroscopic data obtained earlier for these systems, which are interpreted in terms of the presence of surface-related iron(III)-containing forms featured by essentially higher quadrupole splitting values. The surface analysis data presented are also discussed in connection with the high electrocatalytic activity of composite nickel-iron hydroxide systems towards anodic oxygen evolution in alkaline electrolytes revealed previously.

Transmission Mossbauer spectra of binary coprecipitated nickel(II)–iron(III) hydroxides (molar ratios 9:1 and 4:1) are analysed and compared with the results of AES analyses of their surface composition. A correlation between the data of the above techniques has been noted, indicating a redistribution of the components (i.e. Ni and Fe) in the surface layers, the latter being enriched with iron(III) as compared to the bulk. The results obtained are in good agreement with an excellent electrocatalytic activity of composite Ni–Fe hydroxide systems towards anodic oxygen evolution in alkaline electrolytes, taking into account a distinctly exhibited ‘surface’ electrocatalytic effect of ferric species on the nickel hydroxide electrode in the anodic oxygen evolution process revealed previously. Transmission Mossbauer spectroscopy has been shown to be a highly effective probe sensitive both to various bulk and surface effects related to the solid phases studied.

Mossbauer parameters for ferric hydroxo forms in alkaline matrices, ferric oxyhydroxide and its admixture at the nickel hydroxide electrode surface as well as for ferric hydroxy species adsorbed at nickel hydroxides form strongly alkaline solutions have been compared. The feasibility of transmission Mossbauer spectra acquisition for the adsorbed species in the presence of dissolved forms (e.g. in situ) us based on the principle that the latter give no Mossbauer effect at room temperature owing to their diffusional mobility, while the former, being fixed at the adsorbent surface, have been shown to exhibit a sufficient resonant adsorption

Transmission Mossbauer spectra of soluble ferric hydroxo species fixed at the surface of Ni(II) hydroxide and higher Ni oxyhydroxides as a result of their adsorption from concentrated alkaline aqueous solutions at ambient temperature have been obtained. A relatively high adsorption rate and its reversibility have been confirmed; parameters of the Mossbauer spectra obtained provide evidence that the adsorbed ferric species retain their hydroxo coordination while its symmetry is being distorted, which is considered to proceed with partial loss of water molecules of the outer (second) coordination sphere. Specimen treatment in water leads to destruction of the initial coordination, the bonding strength of the resulting ferric particles to the adsorbent surface being significantly increased. The role of the surface phase non-homogeneity of higher Ni oxyhydroxide has also been outlined.

57Fe Mossbauer spectroscopy was used to study the uptake and distribution of iron in the root of cucumber plants grown in iron-deficient modified Hoagland nutrient solution and put into iron-containing solution with 10 μM Fe citrate enriched with 57Fe (90%) only before harvesting. The Mossbauer spectra of the frozen roots exhibited two Fe3+ components with typical average Mossbauer parameters of δ = 0.5 mm s−1, Δ = 0.46 mm s−1 and δ = 0.5 mm s−1, Δ = 1.2 mm s−1 at 78 K and the presence of an Fe2+ doublet, assigned to the ferrous hexaaqua complex. This finding gives a direct evidence for the existence of Fe2+ ions produced via root-associated reduction according to the mechanism proposed for iron uptake for dicotyledonous plants. Monotonous changes in the relative content of the components were found with the time period of iron supply. The Mossbauer results are interpreted in terms of iron uptake and transport through the cell wall and membranes.

Comparative Auger electron spectroscopic data on the surface composition and depth profiling for beta-Ni(OH)(2) and Ni(OH)(2) coprecipitated with iron(III) (3 at.%) are given showing a non-uniform distribution of the latter. They are considered together with their structural characteristics obtained using Fourier transform infrared spectroscopy (both samples) and Mössbauer spectroscopy (Fe-containing sample). The results obtained provide an explanation for the specific behaviour of the Fe(III)-doped nickel(II) hydroxide in heterogeneous processes (adsorption, electrocatalysis).

Interaction of cobalt(II) at micromolar concentrations with live cells of the plant-growth-promoting rhizobacterium Azospirillum brasilense (strain Sp245) and further transformations of the metal cation were monitored using 57Co emission Mössbauer spectroscopy (EMS). Cell suspensions of the bacterial culture (2.4 x 10(8) cells ml(-1)) were doped with radioactive 57CoCl2 (1 mCi; final concentration 2 x 10(-6) M 57Co2+), kept under physiological conditions for various periods of time (from 2 min up to 1 hour) and then rapidly frozen in liquid nitrogen. Analysis of emission Mössbauer spectra of the frozen aqueous suspensions of the bacterial cell samples shows that the primary absorption of cobalt(II) at micromolar concentrations by the bacterial cells is rapid and virtually complete, giving at least two major forms of cobalt(II) species bound to the cells. Within an hour, the metal is involved in further metabolic transformations reflected by changes occurring in the spectra. The Mössbauer parameters calculated from the EMS data by statistical treatment were different for suspensions of live and dead (thermally killed) bacterial cells that had been in contact with 57Co2+ for 1 h, as well as for the cell-free culture medium containing the same concentration of 57Co2+. Chemical after-effects of the nuclear transition (57Co --5 7Fe), which provide additional information on the chemical environment of metal ions, are also considered. The data presented demonstrate that EMS is a valuable tool for monitoring the chemical state of cobalt species in biological matter providing information at the atomic level in the course of its uptake and/or metabolic transformations.

57Co emission Mössbauer spectroscopy (EMS) allows the chemical state of cobalt, as influenced by its coordination environment, to be monitored in biological samples at its physiological (trace) concentrations. To draw attention to EMS as a valuable tool for speciation of cobalt in biocomplexes, the process of cobalt(II) metabolism in cells of the plant growth-promoting rhizobacterium Azospirillum brasilense Sp245 was investigated using EMS of 57CoII-doped bacterial cells. EMS measurements also showed 57CoII-activated glutamine synthetase (GS, a key enzyme of nitrogen metabolism, isolated from this bacterium) to have two different cobalt(II) forms at its active sites, in agreement with data available on other bacterial GSs. Chemical after-effects following electron capture by the nucleus of the parent 57CoII during the 57Co--57Fe transition, which contribute to the formation of a stabilised daughter 57FeIII component along with the nucleogenic 57FeII forms, are also briefly considered.

Fourier transform infrared (FTIR) spectroscopic measurements, performed in the far-IR region (200-800 cm(-1)) on binary coprecipitated Ni(II)-Fe(III) hydroxides (20-60 at.% Fe/(Ni + Fe)), are presented and discussed. Heating the sample containing 60 at.% Fe to 600 degrees C, monitored by differential scanning calorimetry (DSC) measurements, results in the formation of a nickel ferrite structure from its precursor present in binary hydroxides. The role of Fe-to-Ni ratio non-uniformities within the surface and bulk of the binary hydroxide crystallites, as revealed by Mossbauer measurements, Auger electron spectroscopy and depth profiling performed previously, is also discussed with regard to the formation of alpha-Fe2O3 and NiO impurities on thermal treatment.