Effects of electron donors on anaerobic microbial debromination of polybrominated diphenyl ethers (PBDEs)

Polybrominated diphenyl ethers (PBDEs) are a class of widely used flame retardants that have been highly accumulated in sediments. It is reported that microorganisms play an important role in the reductive debromination of PBDEs in anaerobic sediments. However, little is known about the effects of electron donors on the microbial community structure and their debromination capacity in PBDE transformation. In this study, alternate carbon substrates were used as electron donors to enrich the PBDE-debrominating microbial consortia to evaluate the effects of electron donors on PBDE microbial debromination. Decabromodiphenyl ether (BDE-209) was found to be the dominant (more than 50%) PBDEs congener in all consortia, and the percentage of BDE-209 was deceased by 12% (methanol), 11% (ethanol), 8% (acetate), 9% (lactate), 5% (pyruvate), and 11% (no electron donors), while the relative abundances of most lesser-brominated PBDEs increased after 90-day incubation compared to the initial profile of PBDEs. Substantial shifts in the microbial community structure among different amendments were observed based on denaturing gradient gel electrophoresis results. Pseudomonas spp. were identified to be the predominant organisms and the abundances of Band R, which was associated with Pseudomonas sp. SCSWA09, was well correlated with the biodegradation rate of BDE-209. Finally, the microbial community structure was highly correlated with the concentration of deca-BDE, octa-BDE and total nitrogen. These results provide insights into in situ bioremediation of environments contaminated by PBDEs and our understanding of microbial ecology associated with PBDE-debromination.     

Identification and biodegradation efficiency of a newly isolated 2,2′,4,4′-tetrabromodiphenyl ether (BDE-47) aerobic degrading bacterial strain

Polybrominated diphenyl ethers (PBDEs) are bioaccumulative, toxic and persistent, globally distributed organic chemicals in environment. However, very little is known for their aerobic biodegradation. In this research, 2,2′,4,4′-tetrabromodiphenyl ether (BDE-47) was selected as a model congener of PBDEs to study its aerobic biodegradation. A new BDE-47 degrading strain BFR01 identified as Pseudomonas stutzeri was isolated from polluted soil in a former brominated flame retardant production corporation. Stain BFR01 could utilize BDE-47 as a sole source of carbon and energy, and transformed 97.94% of BDE-47 in two weeks; the biodegradation of BDE-47 fitted well with the first-order kinetics, with the first-order kinetics constant of 0.32 d⁻¹. The biodegradation efficiency of stain BFR01 was higher than other reported PBDEs aerobic degrading bacteria. The biodegradation efficiency achieved maximum at pH 7.0 and 40 °C. The presence of additional carbon sources could enhance the biodegradation efficiency of BDE-47 by 1–6%. Furthermore, no lower brominated diphenyl ethers or biphenyl were detected, suggesting that the pathway of BDE-47 biodegradation by strain BFR01 might not be debromination with lower brominated diphenyl ethers as products. This is the first report of aerobic degradation of BDE-47 by P. stutzeri.     

Analysis of debromination of 1,2-dibromoethane by cytochrome P-450-linked hydroxylation systems as observed by bromide electrode

Debromination of 1,2-dibromoethane (DBE) by a rabbit liver microsomal preparation and a reconstituted cytochrome P-450 enzyme system was investigated. The reaction was performed in our newly constructed reaction vessel, in which a bromide electrode was installed. During the reaction, the liberated bromide ion was continuously measured by the bromide electrode, and the amount was recorded. In the microsomal preparation, the DBE-debromination rate per nmol cytochrome P-450 was enhanced by phenobarbital-pretreatment of rabbits compared with the untreated microsomes, whereas it was diminished by 3-methylcholanthrene-pretreatment. The debromination reaction was reconstituted in a purified enzyme system containing phenobarbital-inducible rabbit liver microsomal cytochrome P-450 (P-450PB), NADPH-cytochrome P-450 reductase, and NADPH. The optimum conditions required the presence of dilauroylphosphatidylcholine and cytochrome b5. Cytochrome b5 was found not to be an obligatory component for the DBE-debromination in the reconstituted system, but it stimulated the activity about 3.4-fold. Preincubation of the reconstituted mixture with guinea pig anti-cytochrome P-450PB antiserum markedly inhibited the debromination reaction.     

Assessing the reactivation efficacy of hydroxylamine anion towards VX-inhibited AChE: a computational study

Oximate anions are used as potential reactivating agents for OP-inhibited AChE because of they possess enhanced nucleophilic reactivity due to the α-effect. We have demonstrated the process of reactivating the VX–AChE adduct with formoximate and hydroxylamine anions by applying the DFT approach at the B3LYP/6-311 G(d,p) level of theory. The calculated results suggest that the hydroxylamine anion is more efficient than the formoximate anion at reactivating VX-inhibited AChE. The reaction of formoximate anion and the VX–AChE adduct is a three-step process, while the reaction of hydroxylamine anion with the VX–AChE adduct seems to be a two-step process. The rate-determining step in the process is the initial attack on the VX of the VX–AChE adduct by the nucleophile. The subsequent steps are exergonic in nature. The potential energy surface (PES) for the reaction of the VX–AChE adduct with hydroxylamine anion reveals that the reactivation process is facilitated by the lower free energy of activation (by a factor of 1.7 kcal mol⁻¹) than that of the formoximate anion at the B3LYP/6-311 G(d,p) level of theory. The higher free energy of activation for the reverse reactivation reaction between hydroxylamine anion and the VX–serine adduct further suggests that the hydroxylamine anion is a very good antidote agent for the reactivation process. The activation barriers calculated in solvent using the polarizable continuum model (PCM) for the reactivation of the VX–AChE adduct with hydroxylamine anion were also found to be low. The calculated results suggest that V-series compounds can be more toxic than G-series compounds, which is in accord with earlier experimental observations.     

The counterion influence on cationic lipid-mediated transfection of plasmid DNA

A panel of DOTAP analogs was prepared by altering the anionic counterion that accompanies the trimethylammonium polar domain. The transfection of plasmid DNA into NIH3T3 cells and mouse lung was examined using the counterion analogs. The in vitro transfection activity decreased as follows: DOTAP · bisulfate > trifluoromethanesulfonate ∼ iodide ∼ bromide > dihydrogenphosphate ∼ chloride ∼ acetate > sulfate. A similar activity trend was observed in vivo.     

The effect of ionic liquid media on activity,selectivity and stability of Candida antarctica lipase B in transesterification reactions

Nineteen different 1,3-dialkylimidazolium-based ionic liquids (ILs) were used as reaction media for the synthesis of butyl butyrate by transesterification from vinyl butyrate and 1-butanol catalyzed by Candida antarctica lipase B (CaLB). The reaction was also carried out in hexane as a reference solvent. In all the water-immiscible ILs assayed, the enzymatic activity and selectivity were higher than that obtained in hexane. However, in water-miscible ILs, the activity was lower than in the reference solvent, although they showed >99.99% selectivity. Two solvent properties, hydrophobicity and nucleophilicity, were considered key parameters for analyzing the behavior of CaLB in ILs. In the case of ILs based on the same anion, the synthetic activity was gradually enhanced by increasing cation hydrophobicity. Furthermore, the activity of CaLB was greater in ILs containing anions of lower nucleophilicity. Stability studies indicate that CaLB exhibited greater stability in water-immiscible ILs than in water-miscible ILs.     

Carbon nanotube functionalization with carboxylic derivatives: a DFT study

Chemical functionalization of a single-walled carbon nanotube (CNT) with different carboxylic derivatives including –COOX (X?=?H, CH₃, CH₂NH₂, CH₃Ph, CH₂NO₂, and CH₂CN) has been theoretically investigated in terms of geometric, energetic, and electronic properties. Reaction energies have been calculated to be in the range of ?0.23 to ?7.07 eV. The results reveal that the reaction energy is increased by increasing the electron withdrawing character of the functional groups so that the relative magnitude order is ?CH₂NO₂?>?CH₂CN?>?H?>?CH₂Ph?>?CH₃?>?CH₂NH₂. The chemical functionalization leads to an increase in HOMO/LUMO energy gap of CNT by about 0.32 to 0.35 eV (except for ?H). LUMO, HOMO, and Fermi level of the CNT are shifted to lower energies especially in the case of ?CH₂NO₂ and _?CH₂CN functional groups. Therefore, it leads to an increment in work function of the tube, impeding the field electron emission.     

Aerobic debromination of deca-BDE: Isolation and characterization of an indigenous isolate from a PBDE contaminated sediment

The environmental safety of decabromodiphenyl ether (deca-BDE) has been the topic of controversial discussions during the recent years. Reductive debromination of deca-BDE in the environment was proved to be a significant source of lower-brominated Polybrominated diphenyl ethers (PBDEs) to the ecosystem. Currently, very little is known about the susceptibility of deca-BDE to aerobic biotransformation. Lysinibacillus fusiformis strain DB-1, an aerobic bacterium capable of debromination of deca-BDE, was isolated from sediments of LianjiangRiver, Guiyu in Guangdong of China. DB-1 can efficiently transform deca-BDE to lower brominated BDEs using carbon sources such as lactate, pyruvate and acetate, respectively. In liquid cultures, free bromide concentration accumulated to 1220 ??g L⁻¹ with 6 mg L⁻¹ of the nominal initial concentration of deca-BDE after 72 h aerobic incubation. The resting cell activity tests showed that debromination of deca-BDE by DB-1 was an aerobic process. This is the first report for biotransformation of deca-BDE by an indigenous bacterium isolated from PBDEs contaminated environment.     

Characterization of radicals arising from oxidation of commercially-important essential oils

Inappropriate use of essential oils may entail risks to human health due to mutational events, carcinogenic effects, genetic damages and sensitizing effect caused by generation of reactive oxygen species. In order to detect radicals that are expected to form during their oxidation, we measured the electron spin resonance (ESR) spectra of a standard reaction mixture (I) containing 25?μM flavin mononucleotide, 0.018% several essential oils (or 0.015% geraniol), 1.9 M acetonitrile, 20?mM phosphate buffer (pH 7.4), 0.1 M α-(4-pyridyl-1-oxide)-N-tert-butylnitrone (4-POBN) and 1.0?mM FeSO₄(NH₄)₂SO₄ irradiated with 436?nm visible light (7.8 J/cm²). The ESR peak heights of the standard reaction mixture (I) of the essential oils increased in the following order: tea tree?>?palmarosa?>geranium?>?clary sage?>?petitgrain?>?lavender?>?bergamot?>?frankincense?>?ravintsara?>?ylang ylang?>?lemongrass?>?niaouli?>?eucalyptus globulus?>?peppermint. The ESR peak height of the standard reaction mixture (I) of geraniol, a main component of palmarosa, was comparable to the one of palmarosa (97?±?19% of palmarosa). Furthermore, high performance liquid chromatography (HPLC)-ESR analyses of the standard reaction mixture (I) of palmarosa and geraniol gave the same peaks. The results suggest that the radicals formed in the standard reaction mixture (I) of palmarosa are derived from geraniol. HPLC-ESR-mass spectrometry analyses detected m/z 294 ions, 4-POBN/5-hydroxy-3-methyl-3-pentenyl radical adducts and m/z 320 ions, 4-POBN/C₇O₂H₉ radical adducts in the standard reaction (I) of geraniol. The 5-hydroxy-3-methyl-3-pentenyl and C₇O₂H₉ radicals may be implicated in the sensitizing effect of palmarosa.     

Inhibition of Membrane-Bound Adenosine Triphosphatase Activity from Nicotiana tabacum L. Leaves with Alkyltrimethylammonium Bromide

Cetyltrimethylammonium bromide (CTAB), a cationic detergent,more effectively inhibited the activity of membrane-bound epidermaladenosine triphosphatase (ATPase) of tobacco (Nicotiana tabacumL. cv. Samsun) leaves than anionic or non-ionic detergents.The inhibition of ATPase activity was highly dependent on thelength of the alkyl chain of alkyltrimethylammonium: CTAB >dodecyltrimethylammonium bromide > n-octyltrimethylammoniumbromide trimethylammonium chloride cetyl bromide, comparedat 10^–4 M. The last three derivatives hardly inhibitedthe activity. CTAB inhibition was equivalent to that due toother cationic detergents, cetylpyridinium bromide and cetylamine, but less than that by gramicidin S and tyrocidine andstronger than that by N,N'-dicyclohexylcarbodiimide and vanadate. These results show that a certain length of the alkyl chain(C_n>12) and the combination of both hydrophobic and chargedgroups of a detergent moiety are indispensable for inhibitingthe membrane-bound epidermal ATPase activity. (Received January 26, 1982; Accepted April 10, 1982)     

Metal-mediated allylation of enzymatically oxidized methyl α-D-galactopyranoside

The C-6 unit of methyl α-d-galactopyranoside was selectively modified by combining enzymatic oxidation with an indium-mediated allylation reaction. The Barbier-Grignard type reaction, where a carbonyl group reacts with an allyl halide, proceeds in aqueous solution, even with water as the only solvent; thus carbohydrates can be modified without the need for drying or protection-deprotection steps. The corresponding homoallyl alcohols are produced in high yields of >90% in the reactions with allyl bromide and cinnamyl chloride. The main products were isolated and characterized by GC-MS and NMR spectroscopy.     

Aluminium speciation and phytotoxicity in alkaline soils

Aim

Highly alkaline soils (pH?>?9.0) may adversely affect agricultural crop productivity. Problems encountered include poor structure and nutrient deficiency. Research based on solution cultures suggests that aluminium (Al) phytotoxicity may occur in soils with pH?>?9.0, but little research has been undertaken on actual soils under controlled conditions. The nature of the Al species responsible and the pH regime of the soils when this occurs are unknown.

Methods

The charge and species of Al responsible for this toxicity was investigated using Zeta Potential measurement, Nuclear Magnetic Resonance (NMR) spectroscopy, Al precipitation characteristics and electrical conductivity as a function of pH. An anion exchange resin was used to evaluate Al availability to plants at alkaline pH. To verify Al phytotoxicity, a pot experiment was performed with plants grown at near neutral and high pH, with and without Al.

Results

The anionic aluminate species of aluminium was ubiquitous at highly alkaline pH, and was the dominant charged species at pH 9.2. Aluminium was phytotoxic at high pH, significantly reducing the stem and root development of field pea test plants over and above that caused by alkalinity alone. The effects of both alkalinity in general and aluminium in particular became noticeable at pH 9.0 and debilitating at pH?>?9.2.

Conclusion

As this corresponds to the pH where aluminate becomes dominant, it is probably responsible for the phytotoxicity.     

Modification of quinone electrochemistry by the proteins in the biological electron transfer chains: examples from photosynthetic reaction centers

Quinones such as ubiquinone are the lipid soluble electron and proton carriers in the membranes of mitochondria, chloroplasts and oxygenic bacteria. Quinones undergo controlled redox reactions bound to specific sites in integral membrane proteins such as the cytochrome bc₁ oxidoreductase. The quinone reactions in bacterial photosynthesis are amongst the best characterized, presenting a model to understand how proteins modulate cofactor chemistry. The free energy of ubiquinone redox reactions in aqueous solution and in the Q_A and Q_B sites of the bacterial photosynthetic reaction centers (RCs) are compared. In the primary Q_A site ubiquinone is reduced only to the anionic semiquinone (Q^•−) while in the secondary Q_B site the product is the doubly reduced, doubly protonated quinol (QH₂). The ways in which the protein modifies the relative energy of each reduced and protonated intermediate are described. For example, the protein stabilizes Q^•− while destabilizing Q⁼ relative to aqueous solution through electrostatic interactions. In addition, kinetic and thermodynamic mechanisms for stabilizing the intermediate semiquinones are compared. Evidence for the protein sequestering anionic compounds by slowing both on and off rates as well as by binding the anion more tightly is reviewed.     

NaBH4 in N-methylpyrrolidone: a safe alternative for hydride displacement reaction

Enhanced reactivity of NaBH4 was observed as a solution in N-methylpyrrolidone (NMP). Thus, a simple protocol for debromination of alkyl bromide and sulfonate is devised with NMP as a key solvent. Also described is a new mixed borohydride system, NaBH4-LiOTf-NMP, which works as an alternative to NaBH3CN for the SN2 type displacement. No reports have ever revealed usefulness of NMP in borohydride reduction.     

A criterion for proper cosolvents used for ionic liquids: The Lewis acidic and basic dual nature of propylene carbonate

Probe of the Lewis acidic property of propylene carbonate (PC) has been initiated by temperature-dependent ¹⁹F chemical shifts recorded for 1-butyl-3-methylimidazolium hexafluorophosphate/PC (BMIPF₆/PC) mixtures. Investigation of hyperconjugative interactions by natural bond orbital method reveals the trend of electron-accepting capabilities from anion, PC > dimethyl sulfoxide (DMSO) > N,N-dimethyl formamide (DMF) > acetonitrile (AN). Comparison of solvent-induced ionic association of BMIPF₆ in the four basic aprotic organic solvents, made by monitoring concentration- and temperature-dependent-ratios of solvent diffusion coefficients to BMI⁺-containing components, D_sol/D_BMI, shows that the extent of original ionic association can be reduced by solvent dilution in the same decreasing order, PC ∼ DMSO > DMF > AN. In DMF and AN, two basic solvents having similar dielectric constants, the greater Lewis acid of DMF accounts for its better ability to dissociate aggregates of BMIPF₆. By replacing ethanol with 2,2,2-trifluoroethanol (TFE), the poor miscibility of BMIPF₆ with ethanol can be greatly enhanced due to the CF₃ group, capable of adapting electron density from anions via hyperconjugative interactions. In BMIPF₆/TFE mixtures, values of measured on ¹⁹F resonances show larger aggregates than measured on ¹H resonances, indicating charged clusters prefer anionic to cationic states. This observation can be explained by varying ionic states concerning with unbalanced charged aggregates between positive and negative ones.     

Some aspects of NaBH(4) reduction in NMP

In our solvent optimization study of NaBH(4) reduction, NMP was found to enhance the reactivity. A chemoselective debromination of the bromide and sulfonates can be attained in the new borohydride reagent system: NaBH(4)-LiOTf-NMP. This mixed system worked as an alternative to NaBH(3)CN and Bu(3)SnH for the S(N)2 type displacement of alkylbromide and sulfonate. Also mentioned is an expedient reduction of an azide group into amine by NaBH(4) in NMP without any additive, which offers a convenient protocol for the direct transformation of halides into amines via azide in one flask. Some examples of other reductions were also presented.     

Development of enzyme-linked immunosorbent assay for determination of polybrominated diphenyl ether BDE-121

Our interests are in the development of immunoassay-based fast scanning methods for persistent organic pollutants. To develop the immunoassay method of polybrominated diphenyl ether (PBDE), a model compound of PBDE, 2,3′,4,5′,6-pentabromodiphenylether (BDE-121), has been chosen to develop its antibody and the competitive indirect enzyme-linked immunosorbent assay (ELISA) is developed. The hapten of BDE-121 containing reactive carboxylic acid was synthesized and conjugated to carrier proteins (bovine serum albumin [BSA] and ovalbumin [OVA]). Anti-BDE-121 polyclonal antibody was then developed in rabbits as a result of immunization with the BDE-121–BSA conjugate. The optimal amount of coating antigen BDE-121–OVA conjugate and the dilution of antiserum needed in the ELISA were determined with the checkerboard method, and the effects of the properties of PBST (phosphate-buffered saline and Tween 20) buffer (pH and salt concentration) and chemical solvent (types and concentrations) on the ELISA were investigated to achieve a rapid robust assay with high sensitivity. Under the optimized conditions, the developed indirect ELISA shows a linear detection range from 1.74 to 84.1 ng/ml, with an IC₅₀ value of 8.07 ng/ml and a detection limit of 0.644 ng/ml. In total, 11 kinds of compounds were tested for calculating the cross-reactivity, which was less than 8% for nearly all of them. Real samples were analyzed by the proposed immunoassay and gas chromatography/mass spectrometry (GC/MS).     

The amino acid sequence of Clostridium pasteurianum iron protein, a component of nitrogenase. II. Cyanogen bromide peptides

A total of 10 cyanogen bromide peptides were isolated from the S-beta-carboxymethyl iron protein of nitrogenase. Purification of these peptides was performed mainly by gel filtration on Sephadex G-50; by ascending paper chromatography using the solvent system of pyridine, isoamyl alcohol, 0.1 M ammonium hydroxide; and also, in some cases, with additional steps such as anion exchange column chromatography on Dowex 1-X2 or ascending paper chromatography in an acidic solvent system or by pyridine precipitation of the cyanogen bromide fragment. Sequenator analyses of three large cyanogen bromide peptides (53 to 72 residues) provided tryptic peptide overlap data for the inner portion of the protein. The cyanogen bromide peptides accounted for all of the 273 amino acid residues which were present in the tryptic peptides isolated from carboxymethyl-iron protein (Tanaka, M., Haniu, M., Yasunobu, K. T., and Mortenson, L. E. (1977) J. Biol. Chem. 252, 7081-7088).     

Characterization of an enrichment culture debrominating tetrabromobisphenol A and optimization of its activity under anaerobic conditions

Aim: To study the effects of incubation conditions on the microbial community structure and activity of a TBBPA‐debrominating enrichment culture composed of bacterial and archaeal species. Methods and Results: The effects of the methanogen inhibitor 2‐bromoethanesulfonate (BES), of the antibiotic ampicillin, of substrate (tetrabromobisphenol A, TBBPA) omission and availability of different electron donors on microbial community structure and activity were examined under anaerobic conditions. Debromination of TBBPA was blocked in the presence of ampicillin, while long‐term incubation with BES resulted in delayed debromination activity. The results suggest that the bacterial species responsible for the debromination of TBBPA, while archaeal species involved in electron donor metabolism. The enrichment culture lost its debromination activity after cultivation for 9 months without TBBPA, concomitantly with the disappearance of two DNA bands in a denaturing gradient gel electrophoresis (DGGE) analysis of 16S rRNA gene fragments corresponding to Pelobacter carbinolicus and Sphaerochaeta sp. TQ1 that were present in the original culture. When butyrate was used as an electron donor, TBBPA debromination activity was attenuated. When acetate was used as the electron donor, no debromination was observed and in addition, there was a decrease in the abundance of the mcrA gene. Conclusions: The results indicate that to maintain a high rate of TBBPA debromination activity, it is essential to preserve the microbial community structure (bacterial and archaeal members) of this culture and supply an electron donor that produces high amounts of hydrogen when fermented. Significance and Impact of the Study: The study provides important information for the management of cultures to be used in bioremediation of TBBPA contaminated sites.