Minimizing losses in bio-electrochemical systems: the road to applications

Bio-electrochemical systems (BESs) enable microbial catalysis of electrochemical reactions. Plain electrical power production combined with wastewater treatment by microbial fuel cells (MFCs) has been the primary application purpose for BESs. However, large-scale power production and a high chemical oxygen demand conversion rates must be achieved at a benchmark cost to make MFCs economical competitive in this context. Recently, a number of valuable oxidation or reduction reactions demonstrating the versatility of BESs have been described. Indeed, BESs can produce hydrogen, bring about denitrification, or reductive dehalogenation. Moreover, BESs also appear to be promising in the field of online biosensors. To effectively apply BESs in practice, both biological and electrochemical losses need to be further minimized. At present, the costs of reactor materials have to be decreased, and the volumetric biocatalyst activity in the systems has to be increased substantially. Furthermore, both the ohmic cell resistance and the pH gradients need to be minimized. In this review, these losses and constraints are discussed from an electrochemical viewpoint. Finally, an overview of potential applications and innovative research lines is given for BESs.     

Long-chain fatty acyl-coenzyme A-induced inhibition of glucokinase in pancreatic islets from rats depleted in long-chain polyunsaturated omega3 fatty acids

The metabolism of D-glucose was recently reported to be impaired in pancreatic islets from second generation rats depleted in long-chain polyunsaturated omega3 fatty acids. Considering the increased clearance of circulating non-esterified fatty acids prevailing in these rats, a possible inhibition of glucokinase in insulin-producing cells by endogenous long-chain fatty acyl-CoA was considered. The present study was mainly aimed at assessing the validity of the latter proposal. The activity of glucokinase in islet homogenates, as judged from the increase in D-glucose phosphorylation rate in response to a rise in the concentration of the hexose represented, in the omega3-depleted rats, was only 81.8 +/- 4.8% (n = 11; p < 0.005) of the paired value recorded in control animals. This coincided with the fact that the inclusion of D-glucose 6-phosphate (3.0 mM) and D-fructose 1-phosphate (1.0 mM) in the assay medium resulted in a lesser fractional decrease of D-glucose phosphorylation in omega3-depleted rats than in control animals. Moreover, whereas palmitoyl-CoA (50 microM) decreased the activity of glucokinase by 38.0 +/- 6.0% (n = 4; p < 0.01) in islet homogenates from normal rats, the CoA ester failed to affect significantly the activity of glucokinase in islet homogenates from omega3-depleted rats. These findings afford direct support for the view that glucokinase is indeed inhibited by endogenous long-chain fatty acyl-CoA in islets from omega3-depleted rats, such an inhibition probably participating to the alteration of D-glucose catabolism prevailing in these islets.     

L-glutamine and palmitate catabolism in pancreatic islets from rats depleted in long-chain polyunsaturated omega 3 fatty acids

The catabolism of D-glucose was recently found to be impaired in pancreatic islets from rats depleted in long-chain polyunsaturated omega3 fatty acids. The specificity of this alteration was now investigated by characterizing the oxidative fate of endogenous nutrients in islets preincubated with either L-[U-14C]glutamine or [U-14C]palmitate and then incubated variously in the absence of D-glucose, presence of the hexose or presence of metabolic poisons. Relative to their radioactive content after preincubation, the production of 14CO2 by islets prelabelled with [U-14C]glutamine was higher in omega3-depleted rats than control animals. The enhancing action of D-glucose upon such production was impaired, however, in the omega3-depleted rats. The net uptake of 14C-palmitate and absolute value for 14CO2 output were both increased in omega3-depleted rats, whilst the ratio between 14CO2 output and islet radioactive content was decreased in the same animals. The inhibition of 14CO2 production by metabolic poisons was comparable in all cases. These results are consistent with recent findings on such items as the availability of endogenous amino acids and uptake of unesterified fatty acids in extrapancreatic sites of the omega3-depleted rats. They also support the view that the alteration of D-glucose metabolism in the islets of the latter animals may be attributable, in part at least, to alteration of glucokinase kinetics by high intracellular acyl-CoA levels.     

Hydroquinone dioxygenase from pseudomonas fluorescens ACB: a novel member of the family of nonheme-iron(II)-dependent dioxygenases

Hydroquinone 1,2-dioxygenase (HQDO), an enzyme involved in the catabolism of 4-hydroxyacetophenone in Pseudomonas fluorescens ACB, was purified to apparent homogeneity. Ligandation with 4-hydroxybenzoate prevented the enzyme from irreversible inactivation. HQDO was activated by iron(II) ions and catalyzed the ring fission of a wide range of hydroquinones to the corresponding 4-hydroxymuconic semialdehydes. HQDO was inactivated by 2,2'-dipyridyl, o-phenanthroline, and hydrogen peroxide and inhibited by phenolic compounds. The inhibition with 4-hydroxybenzoate (K(i) = 14 microM) was competitive with hydroquinone. Online size-exclusion chromatography-mass spectrometry revealed that HQDO is an alpha2beta2 heterotetramer of 112.4 kDa, which is composed of an alpha-subunit of 17.8 kDa and a beta-subunit of 38.3 kDa. Each beta-subunit binds one molecule of 4-hydroxybenzoate and one iron(II) ion. N-terminal sequencing and peptide mapping and sequencing based on matrix-assisted laser desorption ionization--two-stage time of flight analysis established that the HQDO subunits are encoded by neighboring open reading frames (hapC and hapD) of a gene cluster, implicated to be involved in 4-hydroxyacetophenone degradation. HQDO is a novel member of the family of nonheme-iron(II)-dependent dioxygenases. The enzyme shows insignificant sequence identity with known dioxygenases.     

Quantifying Community Dynamics of Nitrifiers in Functionally Stable Reactors

A sequential batch reactor (SBR) and a membrane bioreactor (MBR) were inoculated with the same sludge from a municipal wastewater treatment plant, supplemented with ammonium, and operated in parallel for 84 days. It was investigated whether the functional stability of the nitrification process corresponded with a static ammonia-oxidizing bacterial (AOB) community. The SBR provided complete nitrification during nearly the whole experimental run, whereas the MBR showed a buildup of 0 to 2 mg nitrite-N liter⁻¹ from day 45 until day 84. Based on the denaturing gradient gel electrophoresis profiles, two novel approaches were introduced to characterize and quantify the community dynamics and interspecies abundance ratios: (i) the rate of change [Δ_t(week)] parameter and (ii) the Pareto-Lorenz curve distribution pattern. During the whole sampling period, it was observed that neither of the reactor types maintained a static microbial community and that the SBR evolved more gradually than the MBR, particularly with respect to AOB (i.e., average weekly community changes of 12.6% ± 5.2% for the SBR and 24.6% ± 14.3% for the MBR). Based on the Pareto-Lorenz curves, it was observed that only a small group of AOB species played a numerically dominant role in the nitritation of both reactors, and this was true especially for the MBR. The remaining less dominant species were speculated to constitute a reserve of AOB which can proliferate to replace the dominant species. The value of these parameters in terms of tools to assist the operation of activated-sludge systems is discussed.     

Selected Bacterial Strains Protect Artemia spp. from the Pathogenic Effects of Vibrio proteolyticus CW8T2

In this study Vibrio proteolyticus CW8T2 has been identified as a virulent pathogen for Artemia spp. Its infection route has been visualized with transmission electron microscopy. The pathogen affected microvilli and gut epithelial cells, disrupted epithelial cell junctions, and reached the body cavity, where it devastated cells and tissues. In vivo antagonism tests showed that preemptive colonization of the culture water with nine selected bacterial strains protected Artemia juveniles against the pathogenic effects. Two categories of the selected strains could be distinguished: (i) strains providing total protection, as no mortality occurred 2 days after the experimental infection with V. proteolyticus CW8T2, with strain LVS8 as a representative, and (ii) strains providing partial protection, as significant but not total mortality was observed, with strain LVS2 as a representative. The growth of V. proteolyticus CW8T2 in the culture medium was slowed down in the presence of strains LVS2 and LVS8, but growth suppression was distinctly higher with LVS8 than with LVS2. It was striking that the strains that gave only partial protection against the pathogen in the in vivo antagonism test showed also a restricted capability to colonize the Artemia compared to the strains providing total protection. The in vivo antagonism tests and the filtrate experiments showed that probably no extracellular bacterial compounds were involved in the protective action but that the living cells were required to protect Artemia against V. proteolyticus CW8T2.     

Site-specific glycosylation analysis of the bovine lysosomal alpha-mannosidase

Lysosomal alpha-mannosidase is a broad specificity exoglycosidase involved in the ordered degradation of glycoproteins. The bovine enzyme is used as an important model for understanding the inborn lysosomal storage disorder alpha-mannosidosis. This enzyme of about 1,000 amino acids consists of five peptide chains, namely a- to e-peptides and contains eight N-glycosylation sites. The N(497) glycosylation site of the c-peptide chain is evolutionary conserved among LAMANs and is very important for the maintenance of the lysosomal stability of the enzyme. In this work, relying on an approach based on mass spectrometric techniques in combination with exoglycosidase digestions and chemical derivatizations, we will report the detailed structures of the N-glycans and their distribution within six of the eight N-glycosylation sites of the bovine glycoprotein. The analysis of the PNGase F-released glycans from the bovine LAMAN revealed that the major structures fall into three classes, namely high-mannose-type (Fuc(0-1)Glc(0-1)Man(4-9)GlcNAc(2)), hybrid-type (Gal(0-1)Man(4-5)GlcNAc(4)), and complex-type (Fuc(0-1)Gal(0-2)Man(3)GlcNAc(3-5)) N-glycans, with core fucosylation and bisecting GlcNAc. To investigate the exact structure of the N-glycans at each glycosylation site, the peptide chains of the bovine LAMAN were separated using SDS-PAGE and in-gel deglycosylation. These experiments revealed that the N(497) and N(930) sites, from the c- and e-peptides, contain only high-mannose-type glycans Glc(0-1)Man(5-9)GlcNAc(2), including the evolutionary conserved Glc(1)Man(9)GlcNAc(2) glycan, and Fuc(0-1)Man(3-5)GlcNAc(2), respectively. Therefore, to determine the microheterogeneity within the remaining glycosylation sites, the glycoprotein was reduced, carboxymethylated, and digested with trypsin. The tryptic fragments were then subjected to concanavalin A (Con A) affinity chromatography, and the material bound by Con A-Sepharose was purified using reverse-phase high-performance liquid chromatography (HPLC). The tandem mass spectrometry (ESI-MS/MS) and the MALDI analysis of the PNGase F-digested glycopeptides indicated that (1) N(692) and N(766) sites from the d-peptide chain both bear glycans consisting of high-mannose (Fuc(0-1)Man(3-7)GlcNAc(2)), hybrid (Fuc(0-1) Gal(0-1)Man(4-5)GlcNAc(4)), and complex (Fuc(0-1)Gal(0-2)Man(3)GlcNAc(4-5)) structures; and (2) the N(367) site, from the b-peptide chain, is glycosylated only with high-mannose structures (Fuc(0-1)Man(3-5)GlcNAc(2)). Taking into consideration the data obtained from the analysis of either the in-gel-released glycans from the abc- and c-peptides or the tryptic glycopeptide containing the N(367) site, the N(133) site, from the a-peptide, was shown to be glycosylated with truncated and high-mannose-type (Fuc(0-1)Man(4-5)GlcNAc(2)), complex-type (Fuc(0-1)Gal(0-1)Man(3)GlcNAc(5)), and hybrid-type (Fuc(0-1)Gal(0-1)Man(5)GlcNAc(4)) glycans.     

Anaerobic and complementary treatment of domestic sewage in regions with hot climates--a review

This study presents a literature review on the treatment of domestic sewage in controlled environments having the anaerobic process and specifically the upflow anaerobic sludge blanket (UASB) concept as the core, under natural hot conditions. The UASB process application is however beset by the preponderance of suspended solids, and the paper looks at its optimization via pre- and post-treatments to curb the prevailing problems, in the light of possible discharge and re-use/recycling/resource recovery, leading to efficient environmental protection. Pre-treatment clarification could be done with ferric chloride/polyelectrolyte, so that phosphate precipitates during the process. The pre-treated liquid phase can be submitted to a high rate anaerobic process, using the simple and robust UASB technology. In a subsequent post-treatment step, ammonium can be removed by ion exchange using a zeolite column through which the wastewater percolates after leaving the anaerobic digester. The various stages can also eliminate a large fraction of the pathogens present in the raw wastewater, mainly through the pre-treatment sedimentation and the ion exchange filtration. The sludge produced in the precipitation stage can be stabilized in a conventional anaerobic digester. Integration of the different treatment steps provides a sustainable technology to treat domestic sewage under hot climate conditions.