Diversity and spatial distribution of sediment ammonia-oxidizing crenarchaeota in response to estuarine and environmental gradients in the Changjiang Estuary and East China Sea

Ammonia-oxidizing archaea (AOA) have recently been found to be potentially important in nitrogen cycling in a variety of environments, such as terrestrial soils, wastewater treatment reactors, marine waters and sediments, and especially in estuaries, where high input of anthropogenic nitrogen is often experienced. The sedimentary AOA diversity, community structure and spatial distribution in the Changjiang Estuary and the adjacent East China Sea were studied. Multivariate statistical analysis indicated that the archaeal amoA genotype communities could be clustered according to sampling transects, and the station located in an estuarine mixing zone harboured a distinct AOA community. The distribution of AOA communities correlated significantly with the gradients of surface-water salinity and sediment sorting coefficient. The spatial distribution of putative soil-related AOA in certain sampling stations indicated a strong impact of the Changjiang freshwater discharge on the marine benthic microbial ecosystem. Besides freshwater, nutrients, organic matter and suspended particles, the Changjiang Diluted Water might also contribute to the transport of terrestrial archaea into the seawater and sediments along its flow path.

Recovery of protein from discharged wastewater during the production of chitin

Studies were carried out to optimize the conditions for the recovery of protein. The results showed that pH of 6.00 for wastewater, the dosage of 1% chitosan solution in 1% acetic acid aqueous solution of 2.0 ml for 50 ml wastewater and 1% FeCl3 aqueous solution of 2 ml for 50 ml wastewater, the flocculation time of 4.0 h were the optimal conditions for the recovery of protein. The obtained protein sediment contained abundant amino acids, especially isoleucine, methione and lysine that are absent in other protein resource. (c) 2007 Elsevier Ltd. All rights reserved.

Biosorption and desorption of Cd2+ from wastewater by dehydrated shreds of Cladophora fascicularis

The adsorption and desorption of algae Cladophora fascicularis and their relation with initial Cd2+ concentration, initial pH, and co-existing ions were studied. Adsorption equilibrium and biosorption kinetics were established from batch experiments. The adsorption equilibrium was adequately described by the Langmuir isotherm, and biosorption kinetics was in pseudo-second order model. The experiment on co-existing ions showed that the biosorption capacity of biomass decreased with an increasing concentration of competing ions. Desorption experiments indicated that EDTA was efficient desorbent for recovery from Cd2+. With high capacities of metal biosorption and desorption, the biomass of Cladophora fascicularis is promising as a cost-effective biosorbent for the removal of Cd2+ from wastewater.

Sorption and desorption of lead (II) from wastewater by green algae Cladophora fascicularis

Biosorption is an effective method to remove heavy metals from wastewater. In this work, adsorption features of Cladophora fascicularis were investigated as a function of time, initial pH, initial Pb(II) concentrations, temperature and co-existing ions. Kinetics and equilibria were obtained from batch experiments. The biosorption kinetics followed the pseudo-second order model. Adsorption equilibria were well described by the Langmuir and Freundlich isotherm models. The maximum adsorption capacity was 198.5 mg/g at 298 K and pH 5.0. The adsorption processes were endothermic and the biosorption heat was 29.6 kJ/mol. Desorption experiments indicated that 0.01 mol/L Na(2)EDTA was an efficient desorbent for the recovery of Pb(II) from biomass. IR spectrum analysis suggested amido or hydroxy, C=O and C-O could combine intensively with Pb(II). (C) 2006 Elsevier B.V. All rights reserved.

Synthesis and evaluation of a thiourea-modified chitosan derivative applied for adsorption of Hg(II) from synthetic wastewater

In this work, a thiourea-modified chitosan derivative (TMCD) was synthesized through two steps, O-carboxymethylated first and then modified by a polymeric Schiff's base of thiourea/glutaraldehyde. The adsorption behavior of mercury (II) ions onto TMCD was investigated through batch method. The maximum adsorption capacity for Hg(II) was found to be 6.29 mmol/g at pH 5.0 and both kinetic and thermodynamic parameters of the adsorption process were obtained. The results indicated that adsorption process was spontaneous exothermic reaction and kinetically followed pseudo-second-order model. The adsorption experiments also demonstrated TMCD had high adsorption selectivity towards Hg(II) ions when coexisted with Cu(II), Zn(II), Cd(II) and Ca(II) in solution and it could be easily regenerated and efficiently reused. (C) 2010 Elsevier B.V. All rights reserved.

Determination of aliphatic amines from soil and wastewater of a paper mill by pre-column derivatization using HPLC and tandem mass spectrometry (HPLC-MS/MS)

A pre-column derivatization method for the sensitive determination of aliphatic amines using the labeling reagent 1,2-benzo-3,4-dihydrocarbazole-9-ethyl chloroformate (BCEOC) followed by HPLC with fluorescence detection and APCI/NIS identification in positive-ion mode has been developed. The chromophore of 2-(9-carbazole)-ethyl chloroformate (CEOC) reagent was replaced by the 1,2-benzo-3,4-dihydrocarbazole functional group, which resulted in a sensitive fluorescence derivatizing reagent, BCEOC, that could easily and quickly label amines. Derivatives were stable enough to be efficiently analyzed by HPLC and showed an intense protonated molecular ion corresponding m/z [M + H](+) with APCI/MS in positive-ion mode. The collision induced dissociation of the protonated molecular ion formed characteristic fragment ions at m/z 264.1, m/z 246.0 and m/z 218.1, corresponding to the cleavages of CH2CH2O-CO, CH2CH2-OCO, and N-CH2CH2O bonds. Studies on derivatization conditions demonstrated that excellent derivatization yields close to 100% were observed with a 3 to 4-fold molar reagent excess in acetonitrile solvent, in the presence of borate buffer (pH 9.0) at 40 degrees C for 10 min. In addition, the detection responses for BCEOC derivatives were compared with those obtained with CEOC and FMOC as labeling reagents. The ratios I-BCEOC/I-CEOC and I-BCEOC/I-FMOC were, respectively, 1.40-2.76 and 1.36-2.92 for fluorescence responses (here, I was the relative fluorescence intensity). Separation of the amine derivatives had been optimized on an Eclipse XDB-C-8 column. Detection limits calculated from an 0.10 pmol injection, at a signal-to-noise ratio of 3, were 18.65-38.82 fmol (injection volume 10 mu L for fluorescence detection. The relative standard deviations for intraday determination (n = 6) of standard amine derivatives (50 pmol) were 0.0063-0.037% for retention times and 3.36-6.93% for peak areas. The mean intra-and inter-assay precision for all amines were <5.4% and 5.8%, respectively. The recoveries of amines ranged from 96 to 113%. Excellent linear responses were observed with correlation coefficients of >0.9994. The established method provided a simple and highly sensitive technique for the quantitative analysis of trace amounts of aliphatic amines from biological and natural environmental samples.

Electrochromatographic evaluation of a silica monolith capillary column for separation of basic pharmaceuticals

A silica-based monolithic capillary column was prepared via a sol-gel process. The continuous skeleton and large through-pore structure were characterized by scanning electron microscopy (SEM). The native silica monolith has been successfully employed in the electrochromatographic separation of beta-blockers and alkaloids extracted from traditional Chinese medicines (TCMs). Column efficiencies greater than 250000 plates/m for capillary electrochromatography (CEC) separation of basic compounds were obtained. It was observed that retention of basic pharmaceuticals on the silica monolith was mainly contributed by a cation-exchange mechanism. Other retention mechanisms including reversed-phase and normal-phase mechanisms and electrophoresis of basic compounds also played a role in separation. A comparison of the differences between CEC and capillary zone electrophoresis (CZE) separation was also discussed.

Biomimetic emulators of high potential peroxygenases: Implications in bioremediation and metabolic studies

Nowadays, classical (bio)remediation processes are affected by some economical and environmental drawbacks. These approaches often seem to be inadequate, particularly in the perspective of sustainable green processes. Since immobilized metalloporphines can emulate the active site of peroxidases and peroxygenases, their use in several bioremediation processes has been analyzed in this work. The described catalytic reactions use bioinspired, homogenized or heterogenized, commercial porphines and showed a remarkable ability to catalyze substrates oxidation at the expenses of different oxidants such as Oxone and hydrogen peroxide. The biomimetic catalysts have been also investigated about their peroxidase- and peroxygenase-like catalysis and ability to emulate lignolytic peroxidases action and substrate specificity. The adducts showed a remarkable ability to catalyze veratryl alcohol (widely recognized as a simple model compound of lignin) oxidation at the expenses of H2O2. In the perspective of broadening industrial applications of the described catalysts, the oxidation of several pollutants such as durable textile dyes and inorganic sulfides, has been attempted with quite promising results, and some findings open the way toward industrial scaling-up. Accordingly, the inexpensiveness of the synthesis and the mild operational conditions allow these adducts to be proposed as applicable catalysts also for industrial large-scale processes. Besides, these synthetic models are helpful also to understand the behavior of pharmaceuticals, antifungal drugs in this case, in the environment, and to predict the drug metabolism by cytochromes P450. The biomimetic catalysts, for the studied cases, also proved to be much more efficient than the corresponding enzymes.

Resíduos de medicamentos

Projeto de Pós-Graduação/Dissertação apresentado à Universidade Fernando Pessoa como parte dos requisitos para obtenção do grau de Mestre em Ciências Farmacêuticas

Investigation of polyhydroxyalkanoate production by an activated sludge microbial consortium treating artificial dairy wastewater

Petrochemical plastics/polymers are a common feature of day to day living as they occur in packaging, furniture, mobile phones, computers, construction equipment etc. However, these materials are produced from non-renewable materials and are resistant to microbial degradation in the environment. Considerable research has therefore been carried out into the production of sustainable, biodegradable polymers, amenable to microbial catabolism to CO2 and H2O. A key group of microbial polyesters, widely considered as optimal replacement polymers, are the Polyhydroxyalkaonates (PHAs). Primary research in this area has focused on using recombinant pure cultures to optimise PHA yields, however, despite considerable success, the high costs of pure culture fermentation have thus far hindered the commercial viability of PHAs thus produced. In more recent years work has begun to focus on mixed cultures for the optimisation of PHA production, with waste incorporations offering optimal production cost reductions. The scale of dairy processing in Ireland, and the high organic load wastewaters generated, represent an excellent potential substrate for bioconversion to PHAs in a mixed culture system. The current study sought to investigate the potential for such bioconversion in a laboratory scale biological system and to establish key operational and microbial characteristics of same. Two sequencing batch reactors were set up and operated along the lines of an enhanced biological phosphate removal (EBPR) system, which has PHA accumulation as a key step within repeated rounds of anaerobic/aerobic cycling. Influents to the reactors varied only in the carbon sources provided. Reactor 1 received artificial wastewater with acetate alone, which is known to be readily converted to PHA in the anaerobic step of EBPR. Reactor 2 wastewater influent contained acetate and skim milk to imitate a dairy processing effluent. Chemical monitoring of nutrient remediation within the reactors as continuously applied and EBPR consistent performances observed. Qualitative analysis of the sludge was carried out using fluorescence microscopy with Nile Blue A lipophillic stain and PHA production was confirmed in both reactors. Quantitative analysis via HPLC detection of crotonic acid derivatives revealed the fluorescence to be short chain length Polyhydroxybutyrate, with biomass dry weight accumulations of 11% and 13% being observed in reactors 1 and 2, respectively. Gas Chromatography-Mass Spectrometry for medium chain length methyl ester derivatives revealed the presence of hydroxyoctanoic, -decanoic and -dodecanoic acids in reactor 1. Similar analyses in reactor 2 revealed monomers of 3-hydroxydodecenoic and 3-hydroxytetradecanoic acids. Investigation of the microbial ecology of both reactors as conducted in an attempt to identify key species potentially contributing to reactor performance. Culture dependent investigations indicated that quite different communities were present in both reactors. Reactor 1 isolates demonstrated the following species distributions Pseudomonas (82%), Delftia acidovorans (3%), Acinetobacter sp. (5%) Aminobacter sp., (3%) Bacillus sp. (3%), Thauera sp., (3%) and Cytophaga sp. (3%). Relative species distributions among reactor 2 profiled isolates were more evenly distributed between Pseudoxanthomonas (32%), Thauera sp (24%), Acinetobacter (24%), Citrobacter sp (8%), Lactococcus lactis (5%), Lysinibacillus (5%) and Elizabethkingia (2%). In both reactors Gammaproteobacteria dominated the cultured isolates. Culture independent 16S rRNA gene analyses revealed differing profiles for both reactors. Reactor 1 clone distribution was as follows; Zooglea resiniphila (83%), Zooglea oryzae (2%), Pedobacter composti (5%), Neissericeae sp. (2%) Rhodobacter sp. (2%), Runella defluvii (3%) and Streptococcus sp. (3%). RFLP based species distribution among the reactor 2 clones was as follows; Runella defluvii (50%), Zoogloea oryzae (20%), Flavobacterium sp. (9%), Simplicispira sp. (6%), Uncultured Sphingobacteria sp. (6%), Arcicella (6%) and Leadbetterella bysophila (3%). Betaproteobacteria dominated the 16S rRNA gene clones identified in both reactors. FISH analysis with Nile Blue dual staining resolved these divergent findings, identifying the Betaproteobacteria as dominant PHA accumulators within the reactor sludges, although species/strain specific allocations could not be made. GC analysis of the sludge had indicated the presence of both medium chain length as well short chain length PHAs accumulating in both reactors. In addition the cultured isolates from the reactors had been identified previously as mcl and scl PHA producers, respectively. Characterisations of the PHA monomer profiles of the individual isolates were therefore performed to screen for potential novel scl-mcl PHAs. Nitrogen limitation driven PHA accumulation in E2 minimal media revealed a greater propensity among isoates for mcl-pHA production. HPLC analysis indicated that PHB production was not a major feature of the reactor isolates and this was supported by the low presence of scl phaC1 genes among PCR screened isolates. A high percentage distribution of phaC2 mcl-PHA synthase genes was recorded, with the majority sharing high percentage homology with class II synthases from Pseudomonas sp. The common presence of a phaC2 homologue was not reflected in the production of a common polymer. Considerable variation was noted in both the monomer composition and ratios following GC analysis. While co-polymer production could not be demonstrated, potentially novel synthase substrate specificities were noted which could be exploited further in the future.

Proteomic approaches to environmental stress in mussel Mytilus edulis due to emerging classes of anthropogenic pollutants

Anthropogenic pollutant chemicals pose a major threat to aquatic organisms. There is a need for more research on emerging categories of environmental chemicals such as nanomaterials, endocrine disruptors and pharmaceuticals. Proteomics offers options and advantages for early warning of alterations in environmental quality by detecting sub-lethal changes in sentinel species such as the mussel, Mytilus edulis. This thesis aimed to compare the potential of traditional biomarkers (such as enzyme activity measurement) and newer redox proteomic approaches. Environmental proteomics, especially a redox proteomics toolbox, may be a novel way to study pollutant effects on organisms which can also yield information on risks to human health. In particular, it can probe subtle biochemical changes at sub-lethal concentrations and thus offer novel insights to toxicity mechanisms. In the first instance, the present research involved a field-study in three stations in Cork Harbour, Ireland (Haulbowline, Ringaskiddy and Douglas) compared to an outharbour control site in Bantry Bay, Ireland. Then, further research was carried out to detect effects of anthropogenic pollution on selected chemicals. Diclofenac is an example of veterinary and human pharmaceuticals, an emerging category of chemical pollutants, with potential to cause serious toxicity to non-target organisms. A second chemical used for this study was copper which is a key source of contamination in marine ecosystems. Thirdly, bisphenol A is a major anthropogenic chemical mainly used in polycarbonate plastics manufacturing that is widespread in the environment. It is also suspected to be an endocrine disruptor. Effects on the gill, the principal feeding organ of mussels, were investigated in particular. Effects on digestive gland were also investigated to compare different outcomes from each tissue. Across the three anthropogenic chemicals studied (diclofenac, copper and bisphenol A), only diclofenac exposure did not show any significant difference towards glutathione transferase (GST) responses. Meanwhile, copper and bisphenol A significantly increased GST in gill. Glutathione reductase (GR) enzyme analysis revealed that all three chemicals have significant responses in gill. Catalase activity showed significant differences in digestive gland exposed to diclofenac and gills exposed to bisphenol A. This study focused then on application of redox proteomics; the study of the oxidative modification of proteins, to M. edulis. Thiol proteins were labelled with 5-iodoacetamidofluorescein prior to one-dimensional and two-dimensional electrophoresis. This clearly revealed some similarities on a portion of the redox proteome across chemical exposures indicating where toxicity mechanism may be common and where effects are unique to a single treatment. This thesis documents that proteomics is a robust tool to provide valuable insights into possible mechanisms of toxicity of anthropogenic contaminants in M. edulis. It is concluded that future research should focus on gill tissue, on protein thiols and on key individual proteins discovered in this study such as calreticulin and arginine kinase which have not previously been considered as biomarkers in aquatic toxicology prior to this study.

Entry and competition in generic biologics

Patents for several blockbuster biological products are expected to expire soon. The Food and Drug Administration is examining whether biologies can and should be treated like pharmaceuticals with regard to generics. In contrast with pharmaceuticals, which are manufactured through chemical synthesis, biologies are manufactured through fermentation, a process that is more variable and costly. Regulators might require extensive clinical testing of generic biologies to demonstrate equivalence to the branded product. The focus of the debate on generic biologies has been on legal and health concerns, but there are important economic implications. We combine a theoretical model of generic biologies with regression estimates from generic pharmaceuticals to estimate market entry and prices in the generic biologic market. We find that generic biologies will have high fixed costs from clinical testing and from manufacturing, so there will be less entry than would be expected for generic pharmaceuticals. With fewer generic competitors, generic biologies will be relatively close in price to branded biologies. Policy makers should be prudent in estimating financial benefits of generic biologies for consumers and payers. We also examine possible government strategies to promote generic competition. Copyright © 2007 John Wiley & Sons, Ltd.

ANALYSIS AND SIMULATION OF ENERGY USE AND COST AT A MUNICIPAL WASTEWATER TREATMENT PLANT

The cost of electricity, a major operating cost of municipal wastewater treatment plants, is related to influent flow rate, power price, and power load. With knowledge of inflow and price patterns, plant operators can manage processes to reduce electricity costs. Records of influent flow, power price, and load are evaluated for Blue Plains Advanced Wastewater Treatment Plant. Diurnal and seasonal trends are analyzed. Power usage is broken down among treatment processes. A simulation model of influent pumping, a large power user, is developed. It predicts pump discharge and power usage based on wet-well level. Individual pump characteristics are tested in the plant. The model accurately simulates plant inflow and power use for two pumping stations [R2 = 0.68, 0.93 (inflow), R2 =0.94, 0.91(power)]. Wet-well stage-storage relationship is estimated from data. Time-varying wet-well level is added to the model. A synthetic example demonstrates application in managing pumps to reduce electricity cost.

Precipitation of heavy metals from wastewater using simulated flue gas: sequent additions of fly ash, lime and carbon dioxide

Lime is a preferred precipitant for the removal of heavy metals from industrial wastewater due to its relatively low cost. To reduce heavy metal concentration to an acceptable level for discharge, in this work, fly ash was added as a seed material to enhance lime precipitation and the suspension was exposed to CO2 gas. The fly ash-lime-carbonation treatment increased the particle size of the precipitate and significantly improved sedimentation of sludge and the efficiency of heavy metal removal. The residual concentrations of chromium, copper, lead and zinc in effluents can be reduced to (mg L-1) 0.08, 0.14, 0.03 and 0.45, respectively. Examination of the precipitates by XRD and thermal analysis techniques showed that calcium-heavy metal double hydroxides and carbonates were present. The precipitate agglomerated and hardened naturally, facilitating disposal without the need for additional solidification/stabilization measures prior to landfill. It is suggested that fly ash, lime and CO2, captured directly from flue gas, may have potential as a method for wastewater treatment. This method could allow the ex-situ sequestration of CO2, particularly where flue-gas derived CO2 is available near wastewater treatment facilities. (C) 2009 Elsevier Ltd. All rights reserved.