Metabolism of the 4-Hydroxyphenylpyruvate Dioxygenase Inhibitor, Mesotrione, in Multiple-Herbicide-Resistant Palmer amaranth (Amaranthus palmeri).

Metabolic resistance to the maize-selective, HPPD-inhibiting herbicide, mesotrione, occurs via Phase I ring hydroxylation in resistant waterhemp and Palmer amaranth; however, mesotrione detoxification pathways post-Phase I are unknown. This research aims to (1) evaluate Palmer amaranth populations for mesotrione resistance via survivorship, foliar injury, and aboveground biomass, (2) determine mesotrione metabolism rates in Palmer amaranth populations during a time course, and (3) identify mesotrione metabolites including and beyond Phase I oxidation. The Palmer amaranth populations, SYNR1 and SYNR2, exhibited higher survival rates (100%), aboveground biomass (c.a. 50%), and lower injury (25-30%) following mesotrione treatment than other populations studied. These two populations also metabolized mesotrione 2-fold faster than sensitive populations, PPI1 and PPI2, and rapidly formed 4-OH-mesotrione. Additionally, SYNR1 and SYNR2 formed 5-OH-mesotrione, which is not produced in high abundance in waterhemp or naturally tolerant maize. Metabolite features derived from 4/5-OH-mesotrione and potential Phase II mesotrione-conjugates were detected and characterized by liquid chromatography-mass spectrometry (LCMS).

Light-driven biodegradation of azo dyes by Shewanella decolorationis-CdS biohybrid in wastewater lacking electron donors.

The lack of electron donors prevents the effective degradation of azo dyes by bacteria, which severely limits the practical application of conventional biological treatment. Herein, we innovatively designed a bio-photoelectric reduction degradation system composed of CdS and Shewanella decolorationis, which could effectively degrade amaranth in anaerobic conditions driven by light when electron donors were unavailable. Compared with bare S. decolorationis and S. decolorationis (heat-killed)-CdS biohybrid, S. decolorationis-CdS biohybrid had 39.36-fold and 3.82-fold higher first-order kinetic constants, respectively. The morphology, particle size, elemental composition, crystalline type, photovoltaic properties, and band structure of the nanoparticles synthesized by S. decolorationis were carefully examined and analyzed. Light-driven biodegradation experiments showed that amaranth was degraded by the synergy of CdS and S. decolorationis. Reductive degradation of amaranth by electrons was demonstrated by electron and hole trapping. The effect of potential coexisting contaminants, which might serve as hole scavengers, on the degradation of amaranth was evaluated. Membrane protein inhibition experiments also suggested that NADH dehydrogenase, menaquinone, and cytochrome P450 played an important role in electron transfer between CdS and Shewanella decolorationis. The cyclic conversion of NAD+/NADH was probably the most critical rate-limiting step. Electrochemical measurements suggested that faster electron transfer might facilitate the degradation of amaranth. Our findings might contribute to the degradation of azo dyes in wastewater lacking electron donors and deepen our recognition of the microbe-material interface. KEY POINTS: • A BPRDS was constructed with Shewanella decolorationis and CdS. • Amaranth was effectively degraded by BPRDS in anaerobic conditions driven by light. • NDH, MQ, and CYP450 were involved in electron transfer.

Elucidating the effects of different photoanode materials on electricity generation and dye degradation in a sustainable hybrid system of photocatalytic fuel cell and peroxi-coagulation process.

The hybrid system of photocatalytic fuel cell - peroxi-coagulation (PFC-PC) is a sustainable and green technology to degrade organic pollutants and generate electricity simultaneously. In this study, three different types of photocatalysts: TiO2, ZnO and α-Fe2O3 were immobilized respectively on carbon cloth (CC), and applied as photoanodes in the photocatalytic fuel cell of this hybrid system. Photocatalytic fuel cell was employed to drive a peroxi-coagulation process by generating the external voltage accompanying with degrading organic pollutants under UV light irradiation. The degradation efficiency of Amaranth dye and power output in the hybrid system of PFC-PC were evaluated by applying different photoanode materials fabricated in this study. In addition, the effect of light on the photocurrent of three different photoanode materials was investigated. In the absence of light, the reduction of photocurrent percentage was found to be 69.7%, 17.3% and 93.2% in TiO2/CC, ZnO/CC and α-Fe2O3/CC photoanodes, respectively. A maximum power density (1.17 mWcm-2) and degradation of dye (93.8%) at PFC reactor were achieved by using ZnO/CC as photoanode. However, the different photoanode materials at PFC showed insignificant difference in dye degradation trend in the PC reactor. Meanwhile, the degradation trend of Amaranth at PFC reactor was influenced by the recombination rate, electron mobility and band gap energy of photocatalyst among different photoanode materials.

Degradation of dye wastewater with a photoelectric integration process (MPEC): Microbial fuel cells-assisted dual electrodes thin-film photoelectrocatalytic.

A novel photoelectric integration process (MPEC) was developed to degrade Amaranth. In the MPEC, the output voltage of the microbial fuel cells (MFCs) was used to assist the dual slant-placed electrodes thin-film photocatalytic (PC). With two MFCs connected in series, the MPEC process realized the highest decolorization efficiency. It is close to that of the external bias photoelectrocatalytic (PEC), and 7% higher than that of the self-generated electric field-assisted photoelectrocatalytic (SPEC). The feasibility of MPEC pre-treatment and MFC post-treatment of Amaranth was investigated. The results demonstrated that MPEC pre-treatment of Amaranth could improve its biodegradability. The higher MPEC decolorization efficiency indicated the stronger biodegradability of the obtained intermediates and the higher MFC output voltage. When the MPEC decolorization efficiency was gradually increased to 50%, the removal efficiencies of total Chemical Oxygen Demand (COD) by the MPEC and MFC increased; when the decolorization efficiency was increased above 50%, the removal efficiencies became stable. MPEC enhanced the biodegradability efficiently and was applicable to pre-treat textile wastewater.

Dye removal of AR27 with enhanced degradation and power generation in a microbial fuel cell using bioanode of treated clinoptilolite-modified graphite felt.

This work studied the performance of a laboratory-scale microbial fuel cell (MFC) using a bioanode that consisted of treated clinoptilolite fine powder coated onto graphite felt (TC-MGF). The results were compared with another similar MFC that used a bare graphite felt (BGF) bioanode. The anode surfaces provided active sites for the adhesion of the bacterial consortium (NAR-2) and the biodegradation of mono azo dye C.I. Acid Red 27. As a result, bioelectricity was generated in both MFCs. A 98% decolourisation rate was achieved using the TC-MGF bioanode under a fed-batch operation mode. Maximum power densities for BGF and TC-MGF bioanodes were 458.8 ± 5.0 and 940.3 ± 4.2 mW m-2, respectively. GC-MS analyses showed that the dye was readily degraded in the presence of the TC-MGF bioanode. The MFC using the TC-MGF bioanode showed a stable biofilm with no biomass leached out for more than 300 h operation. In general, MFC performance was substantially improved by the fabricated TC-MGF bioanode. It was also found that the TC-MGF bioanode with the stable biofilm presented the nature of exopolysaccharide (EPS) structure, which is suitable for the biodegradation of the azo dye. In fact, the EPS facilitated the shuttling of electrons to the bioanode for the generation of bioelectricity.

Evaluating role of immobilized periphyton in bioremediation of azo dye amaranth.

The aim of this study was to evaluate the bioremediation capabilities of three kinds of periphyton (i.e. epiphyton, metaphyton and epilithon) immobilized in bioreactors to decolorize and biodegrade the sulphonated azo dye, amaranth. Results showed that periphyton dominated by phyla including Cyanobacteria, Proteobacteria and Bacteroidetes. Complete removal of dye was shown by all the biofilms periphyton (epiphyton showed highest removal efficacy) over a range of initial concentrations (50-500mgL-1) within 84h at pH 7 and 30°C. Biodegradation of amaranth was confirmed through FTIR and HPLC and the biodegradation pathways were detected by GC-MS/MS analysis. The azo bonds in the amaranth were successfully broken by periphyton and amaranth was converted to non-toxic, aliphatic compounds including isobutene, acetyl acetate and ethyl acetate. The results showed the potential application of immobilized periphyton at industrial scale for the removal of azo dyes from wastewater containing azo dye amaranth.

Exploration of binding of C.I. Food Red 9 with pepsin by optical spectroscopic and molecular docking methods.

A comprehensive study of the effects of C.I. Food Red 9 on the conformation and activity of pepsin was performed using multi-spectral methods and molecular docking technique. Fluorescence and circular dichroism spectral analyzes showed that C.I. Food Red 9 binding induced the changes of secondary and tertiary structure of pepsin. The activity experimental results indicated that the activity of pepsin decreased remarkably with the increasing concentration of C.I. Food Red 9. Multi non-covalent interactions including hydrogen bonds, hydrophobic, and electrostatic forces played important roles in the complex formation between C.I. Food Red 9 and pepsin. The binding constants of pepsin with C.I. Food Red 9 were (1.21±0.036)×10(4) L mol(-1) (298 K) and (1.05±0.043)×10(4) L mol(-1) (310 K). Moreover, the putative binding site of C.I. Food Red 9 on pepsin was near to activity pocket. This study demonstrates that C.I. Food Red 9 could cause some negative effects on pepsin.

Targeting proteins with toxic azo dyes: a microcalorimetric characterization of the interaction of the food colorant amaranth with serum proteins.

The interaction of amaranth with two homologous serum albumins from human and bovine (HSA and BSA) was studied by microcalorimetry. The binding stoichiometry for the complexation of amaranth to both BSA and HSA was around 1, and the equilibrium constants were (5.79 ± 0.07) × 10(5) and (1.76 ± 0.05) × 10(5) M(-1), respectively. The binding reaction to HSA at 298.15 K was driven by a large negative enthalpic contribution and a small but positive entropic contribution, while to BSA, it was entirely enthalpy-driven and the entropic contribution was unfavorable. Parsing of the standard molar Gibbs energy revealed that the complexation was dominated by non-polyelectrolytic forces. Temperature-dependent isothermal titration calorimetry studies revealed that the enthalpic contribution increased and the entropic contribution decreased with the rise in the temperature but the Gibbs energy change remained almost unaltered. Differential scanning calorimetry results revealed that the binding reaction stabilized the serum albumins significantly against thermal unfolding.

H2-dependent azoreduction by Shewanella oneidensis MR-1: involvement of secreted flavins and both [Ni-Fe] and [Fe-Fe] hydrogenases.

In this paper, the hydrogen (H2)-dependent discoloration of azo dye amaranth by Shewanella oneidensis MR-1 was investigated. Experiments with hydrogenase-deficient strains demonstrated that periplasmic [Ni-Fe] hydrogenase (HyaB) and periplasmic [Fe-Fe] hydrogenase (HydA) are both respiratory hydrogenases of dissimilatory azoreduction in S. oneidensis MR-1. These findings suggest that HyaB and HydA can function as uptake hydrogenases that couple the oxidation of H2 to the reduction of amaranth to sustain cellular growth. This constitutes to our knowledge the first report of the involvement of [Fe-Fe] hydrogenase in a bacterial azoreduction process. Assays with respiratory inhibitors indicated that a menaquinone pool and different cytochromes were involved in the azoreduction process. High-performance liquid chromatography analysis revealed that flavin mononucleotide and riboflavin were secreted in culture supernatant by S. oneidensis MR-1 under H2-dependent conditions with concentration of 1.4 and 2.4 µmol g protein(-1), respectively. These endogenous flavins were shown to significantly accelerate the reduction of amaranth at micromolar concentrations acting as electron shuttles between the cell surface and the extracellular azo dye. This work may facilitate a better understanding of the mechanisms of azoreduction by S. oneidensis MR-1 and may have practical applications for microbiological treatments of dye-polluted industrial effluents.

Properties of NAD (P) H azoreductase from alkaliphilic red bacteria Aquiflexum sp. DL6.

Azoreductase plays a key role in bioremediation and biotransformation of azo dyes. It initializes the reduction of azo bond in azo dye metabolism under aerobic or anaerobic conditions. In the present study, we isolated an alkaliphilic red-colored Aquiflexum sp. DL6 bacterial strain and identified by 16S rRNA method. We report nicotinamide adenine dinucleotide and nicotinamide adenine dinucleotide phosphate-dependent azoreductase purified from Aquiflexum sp. DL6 by a combination of ammonium sulfate precipitation and chromatography methods. The azoreductase was purified up to 30-fold with 37 % recovery. The molecular weight was found to be 80 kDa. The optimum activity was observed at pH 7.4 and temperature 60 °C with amaranth azo dye as a substrate. The thermal stability of azoreductase was up to 80 °C. The azoreductase has shown a wide range of substrate specificity, including azo dyes and nitro aromatic compounds. Metal ions have no significant inhibitory action on azoreductase activity. The apparent K m and V max values for amaranth azo dye were 1.11 mM and 30.77 U/mg protein respectively. This NAD (P) H azoreductase represents the first azoreductase to be characterized from alkaliphilic bacteria.

Batch and continuous biodegradation of Amaranth in plain distilled water by P. aeruginosa BCH and toxicological scrutiny using oxidative stress studies.

Bacterium Pseudomonas aeruginosa BCH was able to degrade naphthylaminesulfonic azo dye Amaranth in plain distilled water within 6 h at 50 mg l(-1) dye concentration. Studies were carried out to find the optimum physical conditions and which came out to be pH 7 and temperature 30 °C. Amaranth could also be decolorized at concentration 500 mg l(-1). Presence of Zn and Hg ions could strongly slow down the decolorization process, whereas decolorization progressed rapidly in presence of Mn. Decolorization rate was increased with increasing cell mass. Induction in intracellular and extracellular activities of tyrosinase and NADH-DCIP reductase along with intracellular laccase and veratryl alcohol oxidase indicated their co-ordinate action during dye biodegradation. Up-flow bioreactor studies with alginate immobilized cells proved the capability of strain to degrade Amaranth in continuous process at 20 ml h(-1) flow rate. Various analytical studies viz.--HPLC, HPTLC, and FTIR gave the confirmation that decolorization was due to biodegradation. From GC-MS analysis, various metabolites were detected, and possible degradation pathway was predicted. Toxicity studies carried out with Allium cepa L. through the assessment of various antioxidant enzymes viz. sulphur oxide dismutase, guaiacol peroxidase, and catalase along with estimation of lipid peroxidation and protein oxidation levels conclusively demonstrated that oxidative stress was generated by Amaranth.

Degradation of the synthetic dye amaranth by the fungus Bjerkandera adusta Dec 1: inference of the degradation pathway from an analysis of decolorized products.

We examined the degradation of amaranth, a representative azo dye, by Bjerkandera adusta Dec 1. The degradation products were analyzed by high performance liquid chromatography (HPLC), visible absorbance, and electrospray ionization time-of-flight mass spectroscopy (ESI-TOF-MS). At the primary culture stage (3 days), the probable reaction intermediates were 1-aminonaphthalene-2,3,6-triol, 4-(hydroxyamino) naphthalene-1-ol, and 2-hydroxy-3-[2-(4-sulfophenyl) hydrazinyl] benzenesulfonic acid. After 10 days, the reaction products detected were 4-nitrophenol, phenol, 2-hydroxy-3-nitrobenzenesulfonic acid, 4-nitrobenzene sulfonic acid, and 3,4'-disulfonyl azo benzene, suggesting that no aromatic amines were created. Manganese-dependent peroxidase activity increased sharply after 3 days culture. Based on these results, we herein propose, for the first time, a degradation pathway for amaranth. Our results suggest that Dec 1 degrades amaranth via the combined activities of peroxidase and hydrolase and reductase action.

Role of iron in azoreduction by resting cells of Shewanella decolorationis S12.

AIM: To investigate the role of soluble and insoluble iron in azoreduction by resting cells of Shewanella decolorationis S12. METHODS AND RESULTS: A series of analytical experiments were carried out. Results showed that insoluble Fe(2) O(3) all delayed the reduction of amaranth but did not inhibit it. Adsorption to Fe(2) O(3) particles by the bacterial cell surface could be the reason leading to the delay in azoreduction. For the soluble iron, an important finding was that azoreduction activities were inhibited by soluble iron in high concentration because of its higher redox potential, and the inhibition was strengthened when the electron donor supply was insufficient. However, activities of azoreduction could be enhanced by low concentration of soluble iron. This stimulating effect was because of the electron transfer but not the cell growth. CONCLUSIONS: The effects of iron on azoreduction by the resting cells depended on the solubility and concentration of the iron compounds, which was different from what was observed by the growing cells in the previous studies. SIGNIFICANCE AND IMPACT OF THE STUDY: This study has both theoretical significance in the microbial physiology and practical significance in the bioremediation of azo dyes-contaminated environment.

Energy generation coupled to azoreduction by membranous vesicles from Shewanella decolorationis S12.

Previous studies have demonstrated that Shewanella decolorationis S12 can grow on the azo compound amaranth as the sole electron acceptor. Thus, to explore the mechanism of energy generation in this metabolism, membranous vesicles (MVs) were prepared and the mechanism of energy generation investigated. The membrane, which was fragmentized during preparation, automatically formed vesicles ranging from 37.5-112.5 nm in diameter under electron micrograph observation. Energy was conserved when coupling the azoreduction by the MVs of an azo compound or Fe(III) as the sole electron acceptor with H2, formate, or lactate as the electron donor. The amaranth reduction by the vesicles was found to be inhibited by specific respiratory inhibitors, including Cu(2+) ions, dicumarol, stigmatellin, and metyrapone, indicating that the azoreduction was indeed a respiration reaction. This finding was further confirmed by the fact that the ATP synthesis was repressed by the ATPase inhibitor N,N'-dicyclohexylcarbodiimide (DCCD). Therefore, this study offers solid evidence of a mechanism of microbial dissimilatory azoreduction on a subcell level.

Decoloration of Amaranth by the white-rot fungus Trametes versicolor. Part I. Statistical analysis.

The white-rot fungus Trametes versicolor decolorized the mono-azo-substituted naphthalenic dye Amaranth. The relationship between the amount of enzymes present in the system and the efficiency of the decoloration process was investigated. The two responses used to quantify the process of decoloration (i.e., initial decoloration rate, v0, and the percent concentration of dye decolorized in 1 h, %c) were correlated with the amount of three enzymes considered for the study (lignin peroxidase, manganese peroxidase, and laccase) and analyzed through stepwise regression analysis (forward, backward, and mixed). The results of the correlation analysis and those of the regression analysis indicated that lignin peroxidase is the enzyme having the greatest influence on the two responses.

Decoloration of Amaranth by the white-rot fungus Trametes versicolor. Part II. Verification study.

The involvement of lignin peroxidase (LiP) in the decoloration of the mono-azo substituted napthalenic dye Amaranth was investigated with pure enzymes and whole cultures of Trametes versicolor. The verification study confirmed that LiP has a direct influence on the initial decoloration rate and showed that another enzyme, which does not need hydrogen peroxide to function and is not a laccase, also plays a role during decoloration. These results confirm the results of a previous statistical analysis. Furthermore, the fungal mycelium affects the performance of the decoloration process.

Reduction and partial degradation mechanisms of naphthylaminesulfonic azo dye amaranth by Shewanella decolorationis S12.

Reduction and biodegradation mechanisms of naphthylaminesulfonic azo dye amaranth using a newly isolated Shewanella decolorationis strain S12 were investigated. Under anaerobic conditions, amaranth was reduced by strain S12, and a stoichiometric amount of two reduction products RP-1 and RP-2 were generated. UV/visible spectrophotometric and high performance liquid chromatography (HPLC) analysis indicated that RP-1 and RP-2 were 1-aminenaphthylene -4-sulfonic acid and 1-aminenaphthylene-2-hydroxy-3, 6-disulfonic acid. The result strongly supports a mechanism of azo dye reduction by the process via the reductive cleavage of the azo bond to form corresponding aromatic amines. The result of HPLC analyses revealed that these aromatic amines were not able to be mineralized by strain S12 under anaerobic conditions. But after re-aeration of the decolorized culture, RP-2 was mineralized completely by this microorganism, but the consumption of RP-1 was not observed. Ames test showed that amaranth had mutagenic but no cytotoxic potential. The mutagenic potential was relieved after the anaerobic treatment with strain S12 as the mutagenic effect of the two reduction products from amaranth was not detected by Ames test. Thus, the ability of strain S12 to reduce and partially mineralize the naphthylaminesulfonic azo dye efficiently was demonstrated, which can potentially be used to biodegrade and detoxify wastewater containing azo dyes using an alternating anaerobic/aerobic treatment procedure.

Functional consequences of blood clotting in insects.

Recent in vitro studies have revealed several important aspects of the biochemical and cellular processes involved in insect blood clotting. However, in vivo empirical studies of the functional consequences of clotting are lacking, despite the role of coagulation in wound-healing, preventing infection, and its homology with vertebrate wound repair. Here we present results of the in vivo effects of haemolymph coagulation and its consequences on the spatial disposition of immune activity, in the American cockroach Periplaneta americana. Our results demonstrate that clotting: (1) localises immune effectors in the vicinity of a breach of the cuticle; (2) restricts the spread of invasive particles across the haemocoel, and (3) is greater when wounding is associated with non-self. Our results demonstrate that haemolymph coagulation has major functional consequences, the most important of which is the compartmentalisation of the open haemocoel.

Respiration and growth of Shewanella decolorationis S12 with an Azo compound as the sole electron acceptor.

The ability of Shewanella decolorationis S12 to obtain energy for growth by coupling the oxidation of various electron donors to dissimilatory azoreduction was investigated. This microorganism can reduce a variety of azo dyes by use of formate, lactate, pyruvate, or H(2) as the electron donor. Furthermore, strain S12 grew to a maximal density of 3.0 x 10(7) cells per ml after compete reduction of 2.0 mM amaranth in a defined medium. This was accompanied by a stoichiometric consumption of 4.0 mM formate over time when amaranth and formate were supplied as the sole electron acceptor and donor, respectively, suggesting that microbial azoreduction is an electron transport process and that this electron transport can yield energy to support growth. Purified membranous, periplasmic, and cytoplasmic fractions from S12 were analyzed, but only the membranous fraction was capable of reducing azo dyes with formate, lactate, pyruvate, or H(2) as the electron donor. The presence of 5 microM Cu(2+) ions, 200 microM dicumarol, 100 microM stigmatellin, and 100 microM metyrapone inhibited anaerobic azoreduction activity by both whole cells and the purified membrane fraction, showing that dehydrogenases, cytochromes, and menaquinone are essential electron transfer components for azoreduction. These results provide evidence that the microbial anaerobic azoreduction is linked to the electron transport chain and suggest that the dissimilatory azoreduction is a form of microbial anaerobic respiration. These findings not only expand the number of potential electron acceptors known for microbial energy conservation but also elucidate the mechanisms of microbial anaerobic azoreduction.

Amaranth decoloration by Trametes versicolor in a rotating biological contacting reactor.

Sequential batch and continuous operation of a rotating biological contacting (RBC) reactor and the effects of dissolved oxygen on the decoloration of amaranth by Trametes versicolor were evaluated. Amaranth belongs to the group of azo dyes which are potential carcinogens and/or mutagens that can be transformed into toxic aryl amines under anaerobic conditions. Cultivation of T. versicolor in a stirred tank reactor was found to be unsuitable for amaranth decoloration due to significant biomass fouling and increase in medium viscosity. Assuming that decoloration follows first-order kinetics, amaranth was decolorized more rapidly when T. versicolor was immobilized on jute twine in a RBC reactor operated either in a sequential batch (k=0.25 h(-1)) or in a continuous (0.051 h(-1)) mode compared to a stirred tank reactor (0.015 h(-1)). Oxygen was found to be essential for decoloration with the highest decoloration rates occurring at oxygen saturation. Although longer retention times resulted in more decoloration when the RBC was operated in the continuous mode (about 33% amaranth decoloration), sequential batch operation gave better results (>95%) under similar nutrient conditions. Our data indicate that the fastest decoloration should occur in the RBC using nitrogen-free Kirk's medium with 1 g/l glucose in sequential batch operation at rotational speeds and/or aeration rates which maintain oxygen saturation in the liquid phase.