Neutrophilic, nitrate-dependent, Fe(II) oxidation by a Dechloromonas species

A species of Dechloromonas, strain UWNR4, was isolated from a nitrate-reducing, enrichment culture obtained from Wisconsin River (USA) sediments. This strain was characterized for anaerobic oxidation of both aqueous and chelated Fe(II) coupled to nitrate reduction at circumneutral pH. Dechloromonas sp. UWNR4 was incubated in anoxic batch reactors in a defined medium containing 4.5–5 mM NO₃ ^?, 6 mM Fe²⁺ and 1–1.8 mM acetate. Strain UWNR4 efficiently oxidized Fe²⁺ with 90 % oxidation of Fe²⁺ after 3 days of incubation. However, oxidation of Fe²⁺ resulted in Fe(III)-hydroxide-encrusted cells and loss of metabolic activity, suggested by inability of the cells to utilize further additions of acetate. In similar experiments with chelated iron (Fe(II)-EDTA), encrusted cells were not produced and further additions of acetate and Fe(II)-EDTA could be oxidized. Although members of the genus Dechloromonas are primarily known as perchlorate and nitrate reducers, our findings suggest that some species could be members of microbial communities influencing iron redox cycling in anoxic, freshwater sediments. Our work using Fe(II)-EDTA also demonstrates that Fe(II) oxidation was microbially catalyzed rather than a result of abiotic oxidation by biogenic NO₂ ^?.     

Treatment of Diesel-Contaminated Soil by Fenton and Persulfate Oxidation with Zero-Valent Iron

The objective of this research is to investigate Fenton and persulfate oxidation with zero-valent iron [Fe(0)] as a batch type ex-situ remediation technology for the treatment of diesel-contaminated soil. Results from batch experiments indicate that Fe(0) is a better catalyst for H₂O₂ and persulfate than Fe²⁺ for the enhancement of Fenton and persulfate oxidation in a batch system. Maximum removal was obtained after 12 h when 1 and 2 g of Fe(0) were added to hydrogen peroxide (250 mg/L) and persulfate (250 mg/L), respectively, in a soil-water system. As the amounts of Fe(0) and persulfate were increased three times at the optimal ratio, the removal of total petroleum hydrocarbon (TPH) was enhanced accordingly. More than 90% of the TPH was removed in 3 h, and the treated soil met the Korean regulation level (500 mg/kg) for TPH. Increased amounts of Fe(0) and hydrogen peroxide (up to 10 g and 1250 mg/L, respectively) also significantly enhanced degradation under the optimal conditions. The results of our study suggest that Fe(0)-assisted Fenton and persulfate oxidation in a batch reactor may be an alternative option to treat diesel-contaminated soil.     

Enhanced growth of Acidovorax sp. strain 2AN during nitrate-dependent Fe(II) oxidation in batch and continuous-flow systems

Microbial nitrate-dependent, Fe(II) oxidation (NDFO) is a ubiquitous biogeochemical process in anoxic sediments. Since most microorganisms that can oxidize Fe(II) with nitrate require an additional organic substrate for growth or sustained Fe(II) oxidation, the energetic benefits of NDFO are unclear. The process may also be self-limiting in batch cultures due to formation of Fe-oxide cell encrustations. We hypothesized that NDFO provides energetic benefits via a mixotrophic physiology in environments where cells encounter very low substrate concentrations, thereby minimizing cell encrustations. Acidovorax sp. strain 2AN was incubated in anoxic batch reactors in a defined medium containing 5 to 6 mM NO₃⁻, 8 to 9 mM Fe²⁺, and 1.5 mM acetate. Almost 90% of the Fe(II) was oxidized within 7 days with concomitant reduction of nitrate and complete consumption of acetate. Batch-grown cells became heavily encrusted with Fe(III) oxyhydroxides, lost motility, and formed aggregates. Encrusted cells could neither oxidize more Fe(II) nor utilize further acetate additions. In similar experiments with chelated iron (Fe(II)-EDTA), encrusted cells were not produced, and further additions of acetate and Fe(II)-EDTA could be oxidized. Experiments using a novel, continuous-flow culture system with low concentrations of substrate, e.g., 100 μM NO₃⁻, 20 μM acetate, and 50 to 250 μM Fe²⁺, showed that the growth yield of Acidovorax sp. strain 2AN was always greater in the presence of Fe(II) than in its absence, and electron microscopy showed that encrustation was minimized. Our results provide evidence that, under environmentally relevant concentrations of substrates, NDFO can enhance growth without the formation of growth-limiting cell encrustations.     

Removal of Cr6+ from groundwater using zero-valence iron in the laboratory

Abstract

In this paper, we describe a series of laboratory experiments which quantify the rate of Cr⁶⁺ reduction by Fe⁰. The main goal of these experiments was to determine the removal efficiency of Cr⁶⁺ by iron. The results indicate that Fe⁰ reduces Cr⁶⁺ to Cr³⁺ under alkaline and slightly acidic conditions. The removal efficiency rises with an increase of the initial concentration of Cr⁶⁺ (1 mg/L to 10 mg/L) when the quantity of Fe⁰ is stable. The removal efficiency increases as the quantity of Fe⁰ is raised when other conditions are constant. The removal efficiency would not be affected by other inorganic ions unless they were present at very high concentrations. When the initial concentration Cr⁶⁺ is 10mg/L and pH is 6.5–7.7, the final concentration of Cr⁶⁺ in effluent is less than 0.05 mg/L and the total Fe is less than 0.3 mg/L in effluent.     

NADH-dependent Fe3+EDTA and oxygen reduction by plasma membrane vesicles from barley roots

Brüggemann, W. and Moog, P. R. 1989. NADH-dependent Fe³⁺EDTA and oxygen reduction by plasma membrane vesicles from barley roots. Biochemical properties of pyridine-dinucleotide-dependent Fe³⁺-EDTA reductase were analysed in purified plasma membranes (PM) from barley (Hordeum vulgare L. cv. Marinka) roots. The enzymatic activity preferred NADH over NADPH as electron donor and it was 3-fold increased in the presence of detergent. The reductase showed a pH optimum of 6.8 and saturable kinetics for NADH with K_m (NADH) of 125 μM and V_max of 143 nmol Fe (mg protein)^-1 min^-1 in the presence of 500 μM Fe³⁺EDTA. For the dependence of the reaction rate on the iron compound, K_m(Fe³⁺EDTA) of 120 μM and V_max of 184 nmol (mg protein)^-1 min^-1 were obtained. The activity was insensitive to superoxide dismutase (SOD; EC 1.15.1.1), catalase (EC 1.11.1.6) and antimycin A, but stimulated by an oxygen-free reaction medium. It could be solubilized by 0.25% (w/v) Triton X-100. The solubilized enzyme revealed one band in native polyacrylamide gel electrophoresis (PAGE) and in isoelectric focussing (IEF) at pl 7.4 by enzyme staining. Major polypeptides with molecular weights of 94, 106, 120 and 205 kDa corresponded to the enzyme-stained band from native PAGE. Analysis of oxygen consumption by the membranes revealed the existence of NADH:CK oxidoreductase activity, which was stimulated by salicylhydroxamic acid (SHAM), chinhydron, Fe³⁺EDTA and Fe³⁺EDTA but not by K₃ [Fe(CN)₆] or K₄[Fe (CN)₆). The stimulating effect of the iron chelates on oxygen consumption was due to Fe²⁺ and could be suppressed by bathophenanthroline disulfonate (BPDS), SOD and p-chloromercurophenylsulfonic acid (PCMS). The results are discussed with respect to the nature of the stimulation.     

SORPTION OF FERROUS AND FERRIC IRON BY EXTRACELLULAR POLYMERIC SUBSTANCES (EPS) FROM ACIDOPHILIC BACTERIA

The sorption of Fe(II) and Fe(III) by extracellular polymeric substances (EPS) of acidophilic bacteria Acidiphilium 3.2Sup(5) and Acidithiobacillus ferrooxidans, harvested from the ecosystem of the Tinto River (Huelva, Spain), was investigated. EPS from mixed cultures of both bacteria (EPS_mixed) and pure cultures of A. 3.2Sup(5) (EPS_pure) were extracted with ethylenediamine tetraacetic acid (EDTA) and were characterized by Fourier-transform infrared (FTIR), electron photoemission (XPS), x-ray diffraction (DRX), and energy dispersive x-ray (EDX) spectroscopy and scanning electron microscopy (SEM). EPS _pure were loaded, in sorption tests, with Fe(II) and Fe(III). The results obtained indicate that the biochemical composition and structure of EPS_mixed was very similar to that of EPS_pure. Besides, results indicate that EPS_mixed adsorbed Fe(II) and Fe(III) by preferential interaction with the carboxyl group, which favored the formation of Fe(II)/Fe(III) oxalates. These species were also formed in EPS_pure loaded with Fe(II)/Fe(III). All this behavior suggested that the sorption of iron by EPS_mixed was similar to sorption of EPS_pure, which fitted the Freundlich model. Thus, the iron uptake of EPS_mixed reached 516.7 ± 23.4 mg Fe/g-EPS at an initial concentration of 2.0 g/L of Fe_total and Fe(II)/Fe(III) ratio of 1.0.     

Enhanced phytoremediation of cadmium and/or benzo(a)pyrene contaminated soil by hyperaccumlator Solanum nigrum L.

The role of same amendment on phytoremediating different level contaminated soils is seldom known. Soil pot culture experiment was used to compare the strengthening roles of cysteine (CY), EDTA, salicylic acid (Sa), and Tween 80 (TW) on hyperaccumulator Solanum nigrum L. phytoremediating higher level of single cadmium (Cd) or Benzo(a)pyrene (BAP) and their co-contaminated soils. Results showed that the Cd capacities (ug pot^?1) in shoots of S. nigrum in the combined treatment T_{0.1EDTA+0.9CY} were the highest for the 5 and 15 mg kg^?1 Cd contaminated soils. When S. nigrum remediating co-contaminated soils with higher levels of Cd and BAP, that is, 5 mg kg^?1 Cd + 1 mg kg^?1 BAP and 15 mg kg^?1 Cd + 2 mg kg^?1 BAP, the treatment T_{0.9CY+0.9Sa+0.3TW} showed the best enhancing remediation role. This results were different with co-contaminated soil with 0.771 mg kg^?1 Cd + 0.024 mg kg^?1 BAP. These results may tell us that the combine used of CY, SA, and TW were more useful for the contaminated soils with higher level of Cd and/or BAP. In the combined treatments of Sa+TW, CY was better than EDTA.     

Electron paramagnetic resonance and Mössbauer spectroscopy of intact mitochondria from respiring <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis>

Mitochondria from respiring cells were isolated under anaerobic conditions. Microscopic images were largely devoid of contaminants, and samples consumed O₂ in an NADH-dependent manner. Protein and metal concentrations of packed mitochondria were determined, as was the percentage of external void volume. Samples were similarly packed into electron paramagnetic resonance tubes, either in the as-isolated state or after exposure to various reagents. Analyses revealed two signals originating from species that could be removed by chelation, including rhombic Fe³⁺ (g = 4.3) and aqueous Mn²⁺ ions (g = 2.00 with Mn-based hyperfine). Three S = 5/2 signals from Fe³⁺ hemes were observed, probably arising from cytochrome c peroxidase and the a₃:Cu_b site of cytochrome c oxidase. Three Fe/S-based signals were observed, with averaged g values of 1.94, 1.90 and 2.01. These probably arise, respectively, from the [Fe₂S₂]⁺ cluster of succinate dehydrogenase, the [Fe₂S₂]⁺ cluster of the Rieske protein of cytochrome bc ₁, and the [Fe₃S₄]⁺ cluster of aconitase, homoaconitase or succinate dehydrogenase. Also observed was a low-intensity isotropic g = 2.00 signal arising from organic-based radicals, and a broad signal with g _ave = 2.02. Mössbauer spectra of intact mitochondria were dominated by signals from Fe₄S₄ clusters (60–85% of Fe). The major feature in as-isolated samples, and in samples treated with ethylenebis(oxyethylenenitrilo)tetraacetic acid, dithionite or O₂, was a quadrupole doublet with ΔE _Q = 1.15 mm/s and δ = 0.45 mm/s, assigned to [Fe₄S₄]²⁺ clusters. Substantial high-spin non-heme Fe²⁺ (up to 20%) and Fe³⁺ (up to 15%) species were observed. The distribution of Fe was qualitatively similar to that suggested by the mitochondrial proteome.     

Spectroscopic description of an unusual protonated ferryl species in the catalase from <Emphasis Type="Italic">Proteus mirabilis</Emphasis> and density functional theory calculations on related models. Consequences for the ferryl protonation state in catalase,peroxidase and chloroperoxidase

The catalase from Proteus mirabilis peroxide-resistant bacteria is one of the most efficient heme-containing catalases. It forms a relatively stable compound II. We were able to prepare samples of compound II from P. mirabilis catalase enriched in ⁵⁷Fe and to study them by spectroscopic methods. Two different forms of compound II, namely, low-pH compound II (LpH II) and high-pH compound II (HpH II), have been characterized by Mössbauer, extended X-ray absorption fine structure (EXAFS) and UV-vis absorption spectroscopies. The proportions of the two forms are pH-dependent and the pH conversion between HpH II and LpH II is irreversible. Considering (1) the Mössbauer parameters evaluated for four related models by density functional theory methods, (2) the existence of two different Fe–O_ferryl bond lengths (1.80 and 1.66 Å) compatible with our EXAFS data and (3) the pH dependence of the α band to β band intensity ratio in the absorption spectra, we attribute the LpH II compound to a protonated ferryl Fe^IV–OH complex (Fe–O approximately 1.80 Å), whereas the HpH II compound corresponds to the classic ferryl Fe^IV=O complex (Fe=O approximately 1.66 Å). The large quadrupole splitting value of LpH II (measured 2.29 mm s^?1 vs. computed 2.15 mm s^?1) compared with that of HpH II (measured 1.47 mm s^?1 vs. computed 1.46 mm s^?1) reflects the protonation of the ferryl group. The relevancy and involvement of such (Fe^IV=O/Fe^IV–OH) species in the reactivity of catalase, peroxidase and chloroperoxidase are discussed.     

Effectiveness of Ethylenediamine-N(o-hydroxyphenylacetic)-N′(p-hydroxyphenylacetic) acid (o,p-EDDHA) to Supply Iron to Plants

The Fe chelate o,p-EDDHA/Fe³⁺, in addition to o,o-EDDHA/Fe³⁺, was found recently to be a component of commercial EDDHA/Fe³⁺ chelates. The European Regulation on fertilisers has included o,p-EDDHA as an authorized chelating agent. The efficacy of o,o-EDDHA/Fe³⁺, o,p-EDDHA/Fe³⁺ and EDTA/Fe³⁺ chelates as Fe sources in plant nutrition was studied. Iron-chelate reductase (FC-R) in young cucumber plants (Cucumis sativus L.) roots reduced o,p-EDDHA/Fe³⁺ faster than o,o-EDDHA/Fe³⁺, EDTA/Fe³⁺ and a commercial source of EDDHA/Fe³⁺. The o,p-EDDHA/Fe³⁺ chelate was also more effective than the o,o-EDDHA/Fe³⁺ in decreasing the severity of Fe-deficiency chlorosis in leaves of young soybean (Glycine max L.) plants grown hydroponically. The o,p-EDDHA ligand was more effective in the short-term than the EDTA and o,o-EDDHA ligands at dissolving Fe from selected Fe minerals and soils. However, the ultimate quantity of dissolve Fe was greatest with the o,o-EDDHA ligand.     

Microbial community analysis of simultaneous ammonium removal and Fe3+ reduction at different influent ammonium concentrations

This study investigates the impacts of influent ammonium concentrations on the microbial community in immobilized heterotrophic ammonium removal system. Klebsiella sp. FC61, the immobilized species, has the ability to perform simultaneous ammonium removal and Fe³⁺ reduction. It was found that average ammonium removal rate decreased from 0.308 to 0.157 mg/L/h, as the influent NH₄ ⁺-N was reduced from 20 to 10 mg/L. Meanwhile, at a total Fe³⁺ concentration of 20 mg/L, the average Fe³⁺ reduction removal efficiency and rate decreased from 44.61% and 0.18 mg/L/h, to 27.10% and 0.11 mg/L/h, respectively. High-throughput sequencing was used to observe microbial communities in bioreactor Samples B1, B2, and B3, after exposure to different influent NH₄ ⁺-N conditions. Results show that higher influent NH₄ ⁺-N concentrations increased microbial richness and diversity and that Klebsiella sp. FC61 play a functional role in the simultaneous removal of NH₄ ⁺-N and Fe³⁺ reduction in bioreactor systems.

     

Simultaneous nitrification and denitrification with different mixed nitrogen loads by a hypothermia aerobic bacterium

Microorganism with simultaneous nitrification and denitrification ability plays a significant role in nitrogen removal process, especially in the eutrophic waters with excessive nitrogen loads. The nitrogen removal capacity of microorganism may suffer from low temperature or nitrite nitrogen source. In this study, a hypothermia aerobic nitrite-denitrifying bacterium, Pseudomonas tolaasii strain Y-11, was selected to determine the simultaneous nitrification and denitrification ability with mixed nitrogen source at 15 °C. The sole nitrogen removal efficiencies of strain Y-11 in simulated wastewater were obtained. After 24 h of incubation at 15 °C, the ammonium nitrogen fell below the detection limit from an initial value of 10.99 mg/L. Approximately 88.0 ± 0.33% of nitrate nitrogen was removed with the initial concentration of 11.78 mg/L and the nitrite nitrogen was not detected with the initial concentration of 10.75 mg/L after 48 h of incubation at 15 °C. Additionally, the simultaneous nitrification and denitrification nitrogen removal ability of P. tolaasii strain Y-11 was evaluated using low concentration of mixed NH₄⁺-N and NO₃^?–N/NO₂^?–N (about 5 mg/L-N each) and high concentration of mixed NH₄⁺–N and NO₃^?–N/NO₂^?–N (about 100 mg/L-N each). There was no nitrite nitrogen accumulation at the time of evaluation. The results demonstrated that P. tolaasii strain Y-11 had higher simultaneous nitrification and denitrification capacity with low concentration of mixed inorganic nitrogen sources and may be applied in low temperature wastewater treatment.     

Light emission from the Fe2+–EDTA–ascorbic acid–H2O2 system strongly enhanced by plant phenolic acids

Oxidative reactions can result in the formation of electronically excited species that undergo radiative decay depending on electronic transition from the excited state to the ground state with subsequent ultra‐weak photon emission (UPE). We investigated the UPE from the Fe²⁺–EDTA (ethylenediaminetetraacetic acid)–AA (ascorbic acid)–H₂O₂ (hydrogen peroxide) system with a multitube luminometer (Peltier‐cooled photon counter, spectral range 380–630 nm). The UPE, of 92.6 μmol/L Fe²⁺, 185.2 μmol/L EDTA, 472 μmol/L AA, 2.6 mmol/L H₂O₂, reached 1217 ± 118 relative light units during 2 min measurement and was about two times higher (P < 0.001) than the UPE of incomplete systems (Fe²⁺–AA–H₂O₂, Fe²⁺–EDTA–H₂O₂, AA–H₂O₂) and medium alone. Substitution of Fe²⁺ with Cr²⁺, Co²⁺, Mn²⁺ or Cu²⁺ as well as of EDTA with EGTA (ethylene glycol‐bis(β‐aminoethyl ether)‐N,N,N′,N′‐tetraacetic acid) or citrate powerfully inhibited UPE. Experiments with scavengers of reactive oxygen species (dimethyl sulfoxide, mannitol, sodium azide, superoxide dismutase) revealed the dependence of UPE only on hydroxyl radicals. Dimethyl sulfoxide at the concentration of 0.74 mmol/L inhibited UPE by 79 ± 4%. Plant phenolics (ferulic, chlorogenic and caffec acids) at the concentration of 870 μmol/L strongly enhanced UPE by 5‐, 13.9‐ and 46.8‐times (P < 0.001), respectively. It is suggested that augmentation of UPE from Fe²⁺–EDTA–AA–H₂O₂ system can be applied for detection of these phytochemicals.     

Reduction of Fe(II)EDTA-NO by a newly isolated <Emphasis Type="Italic">Pseudomonas</Emphasis> sp. strain DN-2 in NO<Subscript><Emphasis Type="Italic">x</Emphasis></Subscript> scrubber solution

Biological reduction of nitric oxide (NO) chelated by ferrous ethylenediaminetetraacetate (Fe(II)EDTA) to N₂ is one of the core processes in a chemical absorption–biological reduction integrated technique for nitrogen oxide (NO _x ) removal from flue gases. A new isolate, identified as Pseudomonas sp. DN-2 by 16S rRNA sequence analysis, was able to reduce Fe(II)EDTA-NO. The specific reduction capacity as measured by NO was up to 4.17 mmol g DCW⁻¹ h⁻¹. Strain DN-2 can simultaneously use glucose and Fe(II)EDTA as electron donors for Fe(II)EDTA-NO reduction. Fe(III)EDTA, the oxidation of Fe(II)EDTA by oxygen, can also serve as electron acceptor by strain DN-2. The interdependency between various chemical species, e.g., Fe(II)EDTA-NO, Fe(II)EDTA, or Fe (III)EDTA, was investigated. Though each complex, e.g., Fe(II)EDTA-NO or Fe(III)EDTA, can be reduced by its own dedicated bacterial strain, strain DN-2 capable of reducing Fe(III)EDTA can enhance the regeneration of Fe(II)EDTA, hence can enlarge NO elimination capacity. Additionally, the inhibition of Fe(II)EDTA-NO on the Fe(III)EDTA reduction has been explored previously. Strain DN-2 is probably one of the major contributors for the continual removal of NO _x due to the high Fe(II)EDTA-NO reduction rate and the ability of Fe(III)EDTA reduction.     

Characteristics of Nitrate Reduction Using Fe (II) as Electron Donor in Activated Sludge

Static experiments were conducted to investigate the effects of environmental factors on nitrate (NO₃^?-N)-removal efficiency, such as NO₃^?-N loading, pH value, C/N ratio and temperature in activated sludge using Fe (II) as electron donor. The results demonstrated that the average denitrification rate increased from 1.25 to 2.23 mg NO₃^?-N/(L·h) with NO₃^?-N loading increased from 30 to 60 mg/L. When pH increased from 7 to 8, the concentration of NO₃^?-N and nitrite (NO₂^?-N) in effluent were all maintained at quite low levels. C/N ratio had little impact on denitrification process, i.e., inorganic carbon (C) source could still be enough for denitrification process with C/N ratio as low as 5. Temperature had a significant effect on the denitrification efficiency, and NO₃^?-N removal efficiency of 92.03%, 96.77%, 97.67% and 98.23% could be obtained with temperature of 25°C, 30°C, 35°C and 40°C, respectively. SEM, XRD and XRF analysis was used to investigate microscopic surface morphology and chemical composition of the denitrifying activated sludge, and mechanism of the nitrate-dependent anaerobic ferrous oxidation (NAFO) bacterias could be explored with this research.     

Role of Thiobacillus thioparus in the biodegradation of carbon disulfide in a biofilter packed with a recycled organic pelletized material

This study reports the biodegradation of carbon disulfide (CS₂) in air biofilters packed with a pelletized mixture of composted manure and sawdust. Experiments were carried out in two lab-scale (1.2 L) biofiltration units. Biofilter B was seeded with activated sludge enriched previously on CS₂-degrading biomass under batch conditions, while biofilter A was left as a negative inoculation control. This inoculum was characterized by an acidic pH and sulfate accumulation, and contained Achromobacter xylosoxidans as the main putative CS₂ biodegrading bacterium. Biofilter operation start-up was unsuccessfully attempted under xerophilic conditions and significant CS₂ elimination was only achieved in biofilter A upon the implementation of an intermittent irrigation regime. Sustained removal efficiencies of 90–100 % at an inlet load of up to 12 g CS₂ m^?3 h^?1 were reached. The CS₂ removal in this biofilter was linked to the presence of the chemolithoautotrophic bacterium Thiobacillus thioparus, known among the relatively small number of species with a reported capacity of growing on CS₂ as the sole energy source. DGGE molecular profiles confirmed that this microbe had become dominant in biofilter A while it was not detected in samples from biofilter B. Conventional biofilters packed with inexpensive organic materials are suited for the treatment of low-strength CS₂ polluted gases (IL <12 g CS₂ m^?3 h^?1), provided that the development of the adequate microorganisms is favored, either upon enrichment or by inoculation. The importance of applying culture-independent techniques for microbial community analysis as a diagnostic tool in the biofiltration of recalcitrant compounds has been highlighted.     

Abatement of Sorbed Crude Oil by Heterogeneous Fenton Process Using A Contaminated Soil Pre-Impregnated with Dissolved Fe(II) and Humic Acid

Dissolved Fe(II) and humic acid (HA) were pre-impregnated into contaminated soil to catalyze hydrogen peroxide to remove crude oil (CO). The effects of parameters such as initial Fe(II), HA and H₂O₂ concentrations on the oxidation of total petroleum hydrocarbon (TPH) were investigated using response surface methodology based on Box–Behnken design. The rate of hydrogen peroxide decomposition is decreased by pre-impregnating with dissolved Fe(II) + HA compared with only pre-impregnated Fe(II) and modified Fenton (MF). Oxygen evolution is the predominant route of hydrogen peroxide decomposition at natural pH. Unlike O₂ evolution, the kinetics of hydroxyl radical (OH^?) production are clearly uncoupled from H₂O₂ decay in these systems. The steady-state hydroxyl radical production rate is higher in the systems with pre-impregnated dissolved Fe(II) and HA, and more significance is the decrease in detectable TPH (70.84% removal efficiency) when soil is pre-impregnated with dissolved 25 mM Fe(II) + 0.7 mg/mL HA, and with the application of 700 mM H₂O₂, possibly due to hydrogen peroxide catalyzed by the iron of this complex (CO-HA–Fe(II)) producing hydroxyl radical in close proximity to the CO. Meanwhile, the removal efficiency of C₂₁–C₃₀ is up to 65.69%, which is 2.6 times higher than that of the MF (25.52%).     

Production and partial characterization of two types of phytase from Aspergillus niger NCIM 563 under submerged fermentation conditions

Novel extracellular phytase was produced by Aspergillus niger NCIM 563 under submerged fermentation conditions at 30 °C in medium containing dextrin and glucose as carbon sources along with sodium nitrate as nitrogen source. Maximum phytase activity (41.47 IU/mL at pH 2.5 and 10.71 IU/mL at pH 4.0) was obtained when dextrin was used as carbon source along with glucose and sodium nitrate as nitrogen source. Nearly 13 times increase in phytase activity was observed when phosphate in the form of KH₂PO₄ (0.004 g/100 mL) was added in the fermentation medium. Physic-chemical properties of partially purified enzyme indicate the possibility of two distinct forms of phytases, Phy I and Phy II. Optimum pH and temperature for Phy I was 2.5 and 60 °C while Phy II was 4.0 and 60 °C, respectively. Phy I was stable in the pH range 1.5–3.5 while Phy II was stable in the wider pH range, 2.0–7.0. Molecular weight of Phy I and Phy II on Sephacryl S-200 was approximately 304 kDa and 183 kDa, respectively. Phy I activity was moderately stimulated in the presence of 1 mM Mg²⁺, Mn²⁺, Ca²⁺ and Fe³⁺ ions and inhibited by Zn²⁺ and Cd²⁺ ions while Phy II activity was moderately stimulated by Fe³⁺ ions and was inhibited by Hg²⁺, Mn²⁺ and Zn²⁺ ions at 1 mM concentration in reaction mixture. The Km for Phy I and II was 3.18 and 0.514 mM while Vmax was 331.16 and 59.47 μmols/min/mg protein, respectively.     

Spin concentration measurements of high-spin (g′ = 4.3) rhombic iron(III) ions in biological samples: theory and application

Electron paramagnetic resonance (EPR) signals at g′ = 4.3 are commonly encountered in biological samples owing to mononuclear high-spin (S = 5/2) Fe³⁺ ions in sites of low symmetry. The present study was undertaken to develop the experimental method and a suitable g′ = 4.3 intensity standard and for accurately quantifying the amount of Fe³⁺ responsible for such signals. By following the work of Aasa and Vänngård (J. Magn. Reson. 19:308–315, 1975), we present equations relating the EPR intensity of S = 5/2 ions to the intensities of S = 1/2 standards more commonly employed in EPR spectrometry. Of the chelates tested, Fe³⁺–EDTA (1:3 ratio) in 1:3 glycerol/water (v/v), pH 2, was found to be an excellent standard for frozen-solution S = 5/2 samples at 77 K. The spin concentrations of Cu²⁺–EDTA and aqua VO²⁺, both S = 1/2 ions, and of Fe³⁺–transferrin, an S = 5/2 ion, were measured against this standard and found to agree within 2.2% of their known metal ion concentrations. Relative standard deviations of ±3.6, ±5.3 and ±2.9% in spin concentration were obtained for the three samples, respectively. The spin concentration determined for Fe³⁺–desferrioxamine of known Fe³⁺ concentration was anomalously low suggesting the presence of EPR-silent multimeric iron species in solution.     

A hypothesis for the basis of the pro-oxidant nature of calcium ions

A new hypothesis describing the role of the redox inactive Ca²⁺ ion in the expression of physiological oxidative damage is described. The hypothesis is based on the optimization of the chelation characteristics of iron complexes for pro-oxidant activity. In a previous investigation it was found that an excess of ligand kinetically hindered the Fenton reaction activity of the Fe^II/IIIEDTA complex (Bobier et al. 2003). EDTA, citrate, NTA, and glutamate were selected as models for the coordination sites likely encountered by mobile iron, i.e. proteins. The optimal [EDTA]:[Fe^III] ratio for Fenton reaction activity as measured by electrocatalytic voltammetry in a solution was found to be 1:1. An excess of EDTA in the amount of 10:1 [ligand]:[metal] suppresses the Fenton reaction activity to nearly the control. It is expected that the physiological coordination characteristics of mobile Fe would have a very large excess of [ligand]:[metal] and thus not be optimized for the Fenton reaction. Introduction of Ca²⁺ in to a ratio of 10:10:1 [Ca²⁺]:[EDTA]:[Fe^III] to the system reinvigorated the Fenton reaction activity to nearly the value of the optimal 1:1 [EDTA]:[Fe^III] complex. The pH distribution diagrams of Ca²⁺ in the presence of EDTA and Fe^II/III indicate that Ca²⁺ has the ability to uptake excess EDTA without displacing either Fe^II of Fe^III from their respective complexed forms. The similarity in the presence for hard ligand sites albeit with a lower binding constant for Ca²⁺ accounts for this action.