Purification and binding analysis of the nitrogen fixation regulatory NifA protein from Azospirillum brasilense

NifA protein activates transcription of nitrogen fixation operons by the alternative sigma54 holoenzyme form of RNA polymerase. This protein binds to a well-defined upstream activator sequence (UAS) located at the -200/-100 position of nif promoters with the consensus motif TGT-N10-ACA. NifA of Azospirillum brasilense was purified in the form of a glutathione-S-transferase (GST)-NifA fusion protein and proteolytic release of GST yielded inactive and partially soluble NifA. However, the purified NifA was able to induce the production of specific anti-A. brasilense NifA-antiserum that recognized NifA from A. brasilense but not from K. pneumoniae. Both GST-NifA and NifA expressed from the E. coli tac promoter are able to activate transcription from the nifHDK promoter but only in an A. brasilense background. In order to investigate the mechanism that regulates NifA binding capacity we have used E. coli total protein extracts expressing A. brasilense nifA in mobility shift assays. DNA fragments carrying the two overlapping, wild-type or mutated UAS motifs present in the nifH promoter region revealed a retarded band of related size. These data show that the binding activity present in the C-terminal domain of A. brasilense NifA protein is still functional even in the presence of oxygen.

Reduction of nitrogen oxide emissions in lime kiln

Different nitrogen oxide removal technologies for rotary lime kiln are studied in this thesis, the main focus being in commercial technologies. Post-combustion methods are investigated in more detail as potential possible NOx removal with combustion methods in rotary lime kiln is more limited or primary methods are already in use. However, secondary methods as NOx scrubber, SNCR or SCR technologies are not listed as the Best Available Technologies defined by European Union. BAT technologies for NOx removal in lime kiln are (1) Optimised combustion and combustion control, (2) Good mixing of fuel and air, (3) Low-NOx burner and (4) Fuel selection/low-N fuel. SNCR method is the most suitable technique for NOx removal in lime kiln when NOx removal from 50 % to 70 % is required in case primary methods are already in use or cannot be applied. In higher removal cases ammonia slip is an issue in SNCR. By using SCR better NOx reduction can be achieved but issues with catalyst materials are expected to arise because of the dust and sulphur dioxide which leads to catalyst poison formation in lower flue gas temperatures. NOx scrubbing has potential when simultaneous NOx and SO2 removal is required. The challenge is that NO cannot be scrubbed directly, but once it is oxidized to NO2 or further scrubbing can be performed as the solubility of NO2 is higher. Commercial installations have not been made regarding SNCR, SCR or NOx scrubbing regarding rotary lime kiln. For SNCR and SCR the closest references come from cement industry.

Different biochemical strategies of two Neotropical fish to cope with the impairment of nitrogen excretion during air exposure

The exposure of fish to air is normally expected to interfere with the nitrogen excretion process. Hoplias malabaricus and Hoplerythrinus unitaeniatus, two teleost species, display distinct behaviors in response to decreases in natural reservoir water levels, although they may employ similar biochemical strategies. To investigate this point, plasma levels of ammonia, urea, uric acid, and the two urea cycle enzymes, ornithine carbamoyl transferase (OCT) and arginase (ARG), as well as glutamine synthetase (GS) were determined for both species after exposure to air. Plasma ammonia increased gradually during exposure to air, but only H. malabaricus showed increased concentrations of urea. Plasma uric acid remained very low in both fish. Enzymatic activities (mean ± SD, µmol min-1 g protein-1) of H. malabaricus showed significant increases (P<0.05, N = 6) in OCT from 0.84 ± 0.05 to 1.42 ± 0.03, in ARG from 8.07 ± 0.47 to 9.97 ± 0.53 and in GS from 1.15 ± 0.03 to 2.39 ± 0.04. The OCT and ARG enzymes remained constant in H. unitaeniatus (N = 6), but GS increased from 1.49 ± 0.02 to 2.06 ± 0.03. Although these species are very closely related and share the same environment, their biochemical strategies in response to exposure to air or to increased plasma ammonia are different.

The biological effects of high-pressure gas on the yeast transcriptome

The aim of the present study was to examine the feasibility of DNA microarray technology in an attempt to construct an evaluation system for determining gas toxicity using high-pressure conditions, as it is well known that pressure increases the concentration of a gas. As a first step, we used yeast (Saccharomyces cerevisiae) as the indicator organism and analyzed the mRNA expression profiles after exposure of yeast cells to nitrogen gas. Nitrogen gas was selected as a negative control since this gas has low toxicity. Yeast DNA microarray analysis revealed induction of genes whose products were localized to the membranes, and of genes that are involved in or contribute to energy production. Furthermore, we found that nitrogen gas significantly affected the transport system in the cells. Interestingly, nitrogen gas also resulted in induction of cold-shock responsive genes. These results suggest the possibility of applying yeast DNA microarray to gas bioassays up to 40 MPa. We therefore think that "bioassays" are ideal for use in environmental control and protection studies.

Effects of pentoxifylline treatment before freezing on motility, viability and acrosome status of poor quality human spermatozoa cryopreserved by the liquid nitrogen vapor method

The objective of the present study was to investigate the effects of the direct addition of pentoxifylline (PF) to the ejaculates of men with poor sperm quality before freezing on post-thaw sperm motility, viability, acrosome integrity, and agonist-induced acrosome reaction. Semen specimens from 16 infertile men with impaired sperm count and motility (oligoasthenozoospermia) were divided into two equal aliquots: one received no treatment (control) while the other was incubated with 5 mM PF (treated). Both aliquots were cryopreserved by the liquid nitrogen vapor method. Motility was assessed according to WHO criteria. Acrosome integrity and spontaneous and calcium ionophore-induced acrosome reactions were assessed with fluorescein isothiocyanate-conjugated peanut agglutinin combined with a supra-vital dye (Hoechst-33258). Cryopreservation impaired sperm motility (percentage reduction: 87.4 (interquartile range, IQ: 70.3-92.9) vs 89.1 (IQ: 72.7-96.0%)), viability (25.9 (IQ: 22.2-29.7) vs 25.6 (IQ: 19.7-40.3%)) and acrosome integrity (18.9 (IQ: 5.4-38.9) vs 26.8 (IQ: 0.0-45.2%)) to the same extent in both treated and control aliquots. However, PF treatment before freezing improved the acrosome reaction to ionophore challenge test scores in cryopreserved spermatozoa (9.7 (IQ: 6.6-19.7) vs 4.8 (IQ: 0.5-6.8%); P = 0.002). These data show that pre-freeze treatment of poor quality human sperm with pentoxifylline did not improve post-thaw motility or viability nor did it prevent acrosomal loss during the freeze-thaw process. However, PF, as used, improved the ability of thawed spermatozoa to undergo the acrosome reaction in response to calcium ionophore. The present data indicate that treatment of poor quality human sperm with PF may enhance post-thaw sperm fertilizing ability.

The selective and non-selective cyclooxygenase inhibitors valdecoxib and piroxicam induce the same postoperative analgesia and control of trismus and swelling after lower third molar removal

We compared the clinical efficacy of orally administered valdecoxib and piroxicam for the prevention of pain, trismus and swelling after removal of horizontally and totally intrabony impacted lower third molars. Twenty-five patients were scheduled to undergo removal of symmetrically positioned lower third molars in two separate appointments. Valdecoxib (40 mg) or piroxicam (20 mg) was administered in a double-blind, randomized and crossed manner for 4 days after the surgical procedures. Objective and subjective parameters were recorded for comparison of postoperative courses. Both agents were effective for postoperative pain relief (N = 19). There was a similar mouth opening at suture removal compared with the preoperative values (86.14 ± 4.36 and 93.12 ± 3.70% of the initial measure for valdecoxib and piroxicam, respectively; ANOVA). There was no significant difference regarding the total amount of rescue medication taken by the patients treated with valdecoxib or piroxicam (173.08 ± 91.21 and 461.54 ± 199.85 mg, respectively; Wilcoxon test). There were no significant differences concerning the swelling observed on the second postoperative day compared to baseline measures (6.15 ± 1.84 and 8.46 ± 2.04 mm for valdecoxib and piroxicam, respectively; ANOVA) or on the seventh postoperative day (1.69 ± 1.61 and 2.23 ± 2.09 mm for valdecoxib and piroxicam, respectively; ANOVA). The cyclooxygenase-2 selective inhibitor valdecoxib is as effective as the non-selective cyclooxygenase inhibitor piroxicam for pain, trismus and swelling control after removal of horizontally and totally intrabony impacted lower third molars.

Cultivation of microalgae Spirulina platensis (Arthrospira platensis) from biological treatment of swine wastewater

The microalgae biomass production from swine wastewater is a possible solution for the environmental impact generated by wastewater discharge into water sources. The biomass can be added to fish feed, which can be used in the formulation of meat products. This work addresses the adaptation of the microalgae Spirulina platensis (Arthrospira platensis) in swine wastewater and the study of the best dilution of the wastewater for maximum biomass production and for removal of Chemical Oxygen Demand (COD), ammonia and phosphorous to the microalgae. The cultivation of Spirulina platensis, strain Paracas presented maximum cellular concentrations and maximum specific growth rates in the wastewater concentration of 5.0 and 8.5%. The highest COD removals occurred with 26.5 and 30.0% of wastewater in the medium. The maximum removal of total phosphorous (41.6%), was with 8.5% of wastewater, which is related to the microalgae growth. The results of Spirulina culture in the swine wastewater demonstrated the possibility of using these microalgae for the COD and phosphorous removal and for biomass production.

Timing of di-ammonium phosphate addition to fermenting chardonnay must : effect on nitrogen utilization, ethyl-carbamate formation, and CAR1 expression by yeast /

The maximum amount of ethyl carbamate (EC), a known animal carcinogen produced by the reaction of urea and ethanol, allowed in alcoholic beverages is regulated by legislation in many countries. Wine yeast produce urea by the metabolism of arginine, the predominant assimilable amino acid in must. This action is due to arginase (encoded by CARl). Regulation of CARl, and other genes in this pathway, is often attributed to a well-documented phenomenon known as nitrogen catabolite repression. The effect of the timing of di-ammonium phosphate (DAP) additions on the nitrogen utilization, regulation of CARl, and EC production was investigated. A correlation was found between the timing of DAP addition and the utilization of nitrogen. When DAP was added earlier in the fermentations, less amino nitrogen and more ammonia nitrogen was sequestered from the media by the cells. It was also seen that early DAP addition led to more total nitrogen being used, with a maximal difference of ~25% between fermentations where no DAP was added versus addition at the start of the fermentation. The effect of the timing ofDAP addition on the expression of CARJ during fermentation was analyzed via northern transfer and the relative levels of CARl expression were determined. The trends in expression can be correlated to the nitrogen data and be used to partially explain differences in EC formation between the treatments. EC was quantified at the end of fermentation by GC/MS. In Montrachet yeast, a significant positive correlation was found between the timing of DAP addition, from early to late, and the final EC concentration m the wine (r = 0.9226). In one of the fermentations, EC levels of 30.5 ppb was foimd when DAP was added at the onset of fermentation. A twofold increase (69.5 ppb) was observed when DAP was added after 75% of the sugars were metabolized. When no DAP was added, the ethyl carbamate levels are comparable at a value of 38 ppb. In contrast, the timing of DAP additions do not affect the level EC produced by the yeast ECU 18 in this manner. The study of additional yeast strains shows that the effect of DAP addition to fermentations is strain dependent. Our results reveal the potential importance of the timing of DAP addition to grape must with respect to EC production, and the regulatory effect of DAP additions on the expression of genes in the pathway for arginine metabolism in certain wine yeast strains.

A biochemical predictor of performance during mesophilic anaerobic fermentation of starch wastewater

The aim of this study was to determine the potential of biochemical parameters, such as enzyme activity and adenosine triphosphate (ATP) levels, as monitors of process performance in the Upflow Anaerobic Sludge Blanket (UASB) reactor utilizing a starch wastewater. The acid and alkaline phosphatase activity and the ATP content of the UASB sludge were measured in response to changes in flow rate and nutrient loading. Conventional parameters of process performance, such as gas production, acetic acid production, COD, phosphorus, nitrogen and suspended solids loadings and % COD removal were also monitored. The response of both biochemical and conventional parameters to changing process conditions was then compared. Alkaline phosphatase activity exhibited the highest activity over the entire study perioda A high suspended solids loading was observed to upset the system in terms of gas production, acetic acid production and % COD removala The initial rate of increase in alkaline phosphatase activity following an increase in loading was four times as great during process upset than under conditions of good performance. The change in enzyme actiVity was also more sensitive to process upset than changes in acetic acid production. The change in ATP content of the sludge with time suggested that enzyme actiVity was changing independently of the actual viable biomass present. The bacterial composition of the anaerobic sludge granules was similar to that of other sludge bed systems, at the light and scanning electron microscope level. Isolated serum bottle cultures produced several acids involved in anaerobic carbohydrate metabolism. The overall performance of the UASB system indicated that higher loadings of soluble nutrients could have been tolerated by the system.

Polystyrene Anchored Vanillin Schiff Base—Complexation and Ion Removal Studies

A set of six new polystyrene anchored metal complexes have been synthesized by the reaction of the metal salt with the polystyrene anchored Schiff base of vanillin. These complexes were characterized by elemental analyses, Fourier transform infrared spectroscopy, diffuse reflectance studies, thermal studies, and magnetic susceptibility measurements. The elemental analyses suggest a metal : ligand ratio of 1 : 2. The ligand is unidentate and coordinates through the azomethine nitrogen. The Mn(II), Fe(III), Co(II), Ni(II), and Cu(II) complexes are all paramagnetic while Zn(II) is diamagnetic. The Cu(II) complex is assigned a square planar structure, while Zn(II) is assigned a tetrahedral structure and Mn(II), Fe(III), Co(II), and Ni(II) are all assigned octahedral geometry. The thermal analyses were done on the ligand and its complexes to reveal their stability. Further, the application of the Schiff base as a chelating resin in ion removal studies was investigated. The polystyrene anchored Schiff base gave 96% efficiency in the removal of Ni(II) from a 20-ppm solution in 15 min, without any interference from ions such as Mn(II), Co(II), Fe(III), Cu(II), Zn(II), U(VI), Na , K , NH4 , Ca2 , Cl , Br , NO3 , NO2 ,and CH3CO2 . The major advantage is that the removal is achieved without altering the pH.

Complexation and ion removal studies of a polystyrene anchored Schiff base

This paper reports the synthesis of a series of six new polystyrene anchored metal complexes of Co(II), Fe(III), Ni(II), Cu(II), Zn(II), and dioxouanium(VI) using the polystyrene anchored Schiff base of 2-nitrobenzaldehyde and the corresponding metal salts. The metal salts used were anhydrous FeCl3, CoCl2 Æ 6H2O, Ni(CH3COO)2 Æ 4H2O, Cu(CH3- COO)2 Æ H2O, Zn(CH3COO)2 Æ 2H2O, and UO2(CH3COO) Æ 2H2O. Physico chemical characterizations have been made from diffuse reflectance and vibrational spectra, elemental analysis, magnetic measurements, and TG studies. The elemental analysis suggest a 1:2 metal:ligand ratio when the complexation has carried out at 70 C for about 12 h reflux. The ligand is monodentate and coordinates through the azomethine nitrogen. The Fe(III), Co(II), Ni(II), and Cu(II) complexes are all paramagnetic whereas Zn(II) and U(VI) are diamagnetic. Zn(II) is assigned a tetrahedral structure, Cu(II) and Co(II) are assigned a square planar structure and Fe(III), Ni(II), and U(VI) are all assigned an octahedral structure. The polystyrene anchored ligand has been developed as an excellent reagent for the removal of Cu(II). Optimum conditions have been developed for the removal of metal ion from solutions by studying the effect of change of concentration of metal ion, ligand, effect of pH, time of reflux, and interference effect of other ions. It was found that within a span of 20 min it is possible to remove 90% of the metal ion from a 30 ppm metal ion solution in the pH range 4–5.5.

Development of bioreactors for nitrifying sewage

Nitrification is the biological oxidation of ammonium, first to nitrite and then to nitrate by two groups of aerobic, chemolithotrophic bacteria belonging to the family Nitrobacteriaceae. The biological nitrification in municipal wastewater treatment is important in those cases were ammonia removal requirement specially exist. In a trickling filter or in an activated sludge system nitrification is rate limiting and thus necessitates longer detention time. The combined carbon oxidation-nitrification processes generally have low population of nitrifiers due to a high ratio of BOD to total nitrogen in the effluent. This necessitates, separate carbon and nitrogen oxidation processes, which thus minimizes wash out ofthe nitrifiers. Therefore, a separate stage nitrification has become essential to achieve faster and efficient removal of ammonia from the wastewater. The present work deals with the development of bio reactor for nitrifying of sewage as the tertiary process so that the treated wastewater can be used for irrigation, algal culture or fish culture

Studies on Advanced Oxidation Processes for the Removal of Acetamiprid from Wastewater

Pollution of water with pesticides has become a threat to the man, material and environment. The pesticides released to the environment reach the water bodies through run off. Industrial wastewater from pesticide manufacturing industries contains pesticides at higher concentration and hence a major source of water pollution. Pesticides create a lot of health and environmental hazards which include diseases like cancer, liver and kidney disorders, reproductive disorders, fatal death, birth defects etc. Conventional wastewater treatment plants based on biological treatment are not efficient to remove these compounds to the desired level. Most of the pesticides are phyto-toxic i.e., they kill the microorganism responsible for the degradation and are recalcitrant in nature. Advanced oxidation process (AOP) is a class of oxidation techniques where hydroxyl radicals are employed for oxidation of pollutants. AOPs have the ability to totally mineralise the organic pollutants to CO2 and water. Different methods are employed for the generation of hydroxyl radicals in AOP systems. Acetamiprid is a neonicotinoid insecticide widely used to control sucking type insects on crops such as leafy vegetables, citrus fruits, pome fruits, grapes, cotton, ornamental flowers. It is now recommended as a substitute for organophosphorous pesticides. Since its use is increasing, its presence is increasingly found in the environment. It has high water solubility and is not easily biodegradable. It has the potential to pollute surface and ground waters. Here, the use of AOPs for the removal of acetamiprid from wastewater has been investigated. Five methods were selected for the study based on literature survey and preliminary experiments conducted. Fenton process, UV treatment, UV/ H2O2 process, photo-Fenton and photocatalysis using TiO2 were selected for study. Undoped TiO2 and TiO2 doped with Cu and Fe were prepared by sol-gel method. Characterisation of the prepared catalysts was done by X-ray diffraction, scanning electron microscope, differential thermal analysis and thermogravimetric analysis. Influence of major operating parameters on the removal of acetamiprid has been investigated. All the experiments were designed using central compoiste design (CCD) of response surface methodology (RSM). Model equations were developed for Fenton, UV/ H2O2, photo-Fenton and photocatalysis for predicting acetamiprid removal and total organic carbon (TOC) removal for different operating conditions. Quality of the models were analysed by statistical methods. Experimental validations were also done to confirm the quality of the models. Optimum conditions obtained by experiment were verified with that obtained using response optimiser. Fenton Process is the simplest and oldest AOP where hydrogen peroxide and iron are employed for the generation of hydroxyl radicals. Influence of H2O2 and Fe2+ on the acetamiprid removal and TOC removal by Fenton process were investigated and it was found that removal increases with increase in H2O2 and Fe2+ concentration. At an initial concentration of 50 mg/L acetamiprid, 200 mg/L H2O2 and 20 mg/L Fe2+ at pH 3 was found to be optimum for acetamiprid removal. For UV treatment effect of pH was studied and it was found that pH has not much effect on the removal rate. Addition of H2O2 to UV process increased the removal rate because of the hydroxyl radical formation due to photolyis of H2O2. An H2O2 concentration of 110 mg/L at pH 6 was found to be optimum for acetamiprid removal. With photo-Fenton drastic reduction in the treatment time was observed with 10 times reduction in the amount of reagents required. H2O2 concentration of 20 mg/L and Fe2+ concentration of 2 mg/L was found to be optimum at pH 3. With TiO2 photocatalysis improvement in the removal rate was noticed compared to UV treatment. Effect of Cu and Fe doping on the photocatalytic activity under UV light was studied and it was observed that Cu doping enhanced the removal rate slightly while Fe doping has decreased the removal rate. Maximum acetamiprid removal was observed for an optimum catalyst loading of 1000 mg/L and Cu concentration of 1 wt%. It was noticed that mineralisation efficiency of the processes is low compared to acetamiprid removal efficiency. This may be due to the presence of stable intermediate compounds formed during degradation Kinetic studies were conducted for all the treatment processes and it was found that all processes follow pseudo-first order kinetics. Kinetic constants were found out from the experimental data for all the processes and half lives were calculated. The rate of reaction was in the order, photo- Fenton>UV/ H2O2>Fenton> TiO2 photocatalysis>UV. Operating cost was calculated for the processes and it was found that photo-Fenton removes the acetamiprid at lowest operating cost in lesser time. A kinetic model was developed for photo-Fenton process using the elementary reaction data and mass balance equations for the species involved in the process. Variation of acetamiprid concentration with time for different H2O2 and Fe2+ concentration at pH 3 can be found out using this model. The model was validated by comparing the simulated concentration profiles with that obtained from experiments. This study established the viability of the selected AOPs for the removal of acetamiprid from wastewater. Of the studied AOPs photo- Fenton gives the highest removal efficiency with lowest operating cost within shortest time.

Sono, Photo and Sonophoto Catalytic Removal of Chemical and Bacterial Pollutants from Wastewater

The term ‘water pollution’ broadly refers to the contamination of water and water bodies (e.g. lakes, rivers, oceans, groundwater etc). Water pollution occurs when pollutants are discharged directly or indirectly into water bodies without adequate treatment to remove the harmful contaminants. This affects not only the plants and organisms living in these bodies of water but also the entire natural biological communities and the biodiversity.Advanced Oxidation Processes (AOPs) have been tested as environment-friendly techniques for the treatment of contaminated water, in view of their ability to convert pollutants into harmless end products. These techniques refer to a set of treatment procedures designed to remove organic or inorganic contaminants in wastewater by oxidation. The contaminants are oxidized by different reagents such as air, oxygen, ozone, and hydrogen peroxide which are introduced in precise, preprogrammed dosages, sequences and combinations under appropriate conditions. The procedure when combined with light in presence of catalyst is known as photocatalysis. When ultrasound (US) is used as the energy source, the process is referred as sonication. Sonication in presence of catalyst is referred as sonocatalysis. Of late, combination of light and sound as energy sources has been tested for the decontamination of wastewater in the presence of suitable catalyst. In this case, the process is referred as sonophotocatalysis. These AOPs are specially advantageous in pollution control and waste water treatment because unlike many other technologies, they do not just transfer the pollutant from one phase to another but completely degrade them into innocuous substances such as CO2 and H2O.

Cereal/legume rotations affect chemical properties and biological activities in two West African soils

A more widespread use of cereal/legume rotations has been suggested as a means to sustainably meet increasing food demands in sub-Saharan West Africa. Enhanced cereal yields following legumes have been attributed to chemical and biological factors such as higher levels of mineral nitrogen (Nmin) and arbuscular mycorrhizae (AM) but also to lower amounts of plant parasitic nematodes. This study was conducted under controlled conditions to examine the relative contribution of AM, plant parasitic nematodes and increased nitrogen (N) and phosphorus (P) availability to cereal/legume rotation effects on two West African soils. Sample soils were taken from field experiments at Gaya (Niger) and Fada (Burkina Faso) supporting continuous cereal and cereal/legume rotation systems and analysed for chemical and biological parameters. Average increases in cereal shoot dry matter (DM) of rotation cereals compared with continuous cereals were 490% at Gaya and 550% at Fada. Shoot P concentration of rotation millet was significantly higher than in continuous millet and P uptake in rotation cereals was on average 62.5-fold higher than in continuous cereals. Rotation rhizosphere soils also had higher pH at both sites. For the Fada soil, large increases in Bray1-P and organic P were observed in bulk and rhizosphere soils. Plant parasitic nematodes in roots of continuous cereals were 60–80-fold higher than in those of rotation cereals. In both cropping systems mycorrhizal infection rates were similar at 37 days after sowing (DAS) but at 57 DAS AM infection was 10–15% higher in rotation sorghum than in continuous sorghum. This study provides strong evidence that cereal/legume rotations can enhance P nutrition of cereals through improved soil chemical P availability and microbiologically increased P uptake.