Inhibition Studies with 2-Bromoethanesulfonate Reveal a Novel Syntrophic Relationship in Anaerobic Oleate Degradation.

Degradation of long-chain fatty acids (LCFAs) in methanogenic environments is a syntrophic process involving the activity of LCFA-degrading bacteria and hydrogen-utilizing methanogens. If methanogens are inhibited, other hydrogen scavengers are needed to achieve complete LCFA degradation. In this work, we developed two different oleate (C18:1 LCFA)-degrading anaerobic enrichment cultures, one methanogenic (ME) and another in which methanogenesis was inhibited (IE). Inhibition of methanogens was attained by adding a solution of 2-bromoethanesulfonate (BrES), which turned out to consist of a mixture of BrES and isethionate. Approximately 5 times faster oleate degradation was accomplished by the IE culture compared with the ME culture. A bacterium closely related to Syntrophomonas zehnderi (99% 16S rRNA gene identity) was the main oleate degrader in both enrichments, in syntrophic relationship with hydrogenotrophic methanogens from the genera Methanobacterium and Methanoculleus (in ME culture) or with a bacterium closely related to Desulfovibrio aminophilus (in IE culture). A Desulfovibrio species was isolated, and its ability to utilize hydrogen was confirmed. This bacterium converted isethionate to acetate and sulfide, with or without hydrogen as electron donor. This bacterium also utilized BrES but only after 3 months of incubation. Our study shows that syntrophic oleate degradation can be coupled to desulfonation.IMPORTANCE In anaerobic treatment of complex wastewater containing fat, oils, and grease, high long-chain fatty acid (LCFA) concentrations may inhibit microbial communities, particularly those of methanogens. Here, we investigated if anaerobic degradation of LCFAs can proceed when methanogens are inhibited and in the absence of typical external electron acceptors, such as nitrate, iron, or sulfate. Inhibition studies were performed with the methanogenic inhibitor 2-bromoethanesulfonate (BrES). We noticed that, after autoclaving, BrES underwent partial hydrolysis and turned out to be a mixture of two sulfonates (BrES and isethionate). We found out that LCFA conversion proceeded faster in the assays where methanogenesis was inhibited, and that it was dependent on the utilization of isethionate. In this study, we report LCFA degradation coupled to desulfonation. Our results also showed that BrES can be utilized by anaerobic bacteria.

Epigenetics in ENS development and Hirschsprung disease.

Hirschsprung disease (HSCR, OMIM 142623) is a neurocristopathy caused by a failure of the enteric nervous system (ENS) progenitors derived from neural crest cells (NCCs), to migrate, proliferate, differentiate or survive to and within the gastrointestinal tract, resulting in aganglionosis in the distal colon. The formation of the ENS is a complex process, which is regulated by a large range of molecules and signalling pathways involving both the NCCs and the intestinal environment. This tightly regulated process needs correct regulation of the expression of ENS specific genes. Alterations in the expression of these genes can have dramatic consequences. Several mechanisms that control the expression of genes have been described, such as DNA modification (epigenetic mechanisms), regulation of transcription (transcription factor, enhancers, repressors and silencers), post-transcriptional regulation (3'UTR and miRNAs) and regulation of translation. In this review, we focus on the epigenetic DNA modifications that have been described so far in the context of the ENS development. Moreover we describe the changes that are found in relation to the onset of HSCR.

ATP5PO levels regulate enteric nervous system development in zebrafish, linking Hirschsprung disease to Down Syndrome.

Hirschsprung disease (HSCR) is a complex genetic disorder characterized by the absence of enteric nervous system (ENS) in the distal region of the intestine. Down Syndrome (DS) patients have a >50-fold higher risk of developing HSCR than the general population, suggesting that overexpression of human chromosome 21 (Hsa21) genes contribute to HSCR etiology. However, identification of responsible genes remains challenging. Here, we describe a genetic screening of potential candidate genes located on Hsa21, using the zebrafish. Candidate genes were located in the DS-HSCR susceptibility region, expressed in the human intestine, were known potential biomarkers for DS prenatal diagnosis, and were present in the zebrafish genome. With this approach, four genes were selected: RCAN1, ITSN1, ATP5PO and SUMO3. However, only overexpression of ATP5PO, coding for a component of the mitochondrial ATPase, led to significant reduction of ENS cells. Paradoxically, in vitro studies showed that overexpression of ATP5PO led to a reduction of ATP5PO protein levels. Impaired neuronal differentiation and reduced mitochondrial ATP production, were also detected in vitro, after overexpression of ATP5PO in a neuroblastoma cell line. Finally, epistasis was observed between ATP5PO and ret, the most important HSCR gene. Taken together, our results identify ATP5PO as the gene responsible for the increased risk of HSCR in DS patients in particular if RET variants are also present, and show that a balanced expression of ATP5PO is required for normal ENS development.

Moorella stamsii sp. nov., a new anaerobic thermophilic hydrogenogenic carboxydotroph isolated from digester sludge.

A novel anaerobic, thermophilic, carbon monoxide-utilizing bacterium, strain E3-O(T), was isolated from anaerobic sludge from a municipal solid waste digester. Cells were straight rods, 0.6-1 µm in diameter and 2-3 µm in length and grew as single cells or in pairs. Cells formed round terminal endospores. The temperature range for growth was 50-70 °C, with an optimum at 65 °C. The pH range for growth was 5.7-8.0, with an optimum at 7.5. Strain E3-O(T) had the ability to ferment various sugars, such as fructose, galactose, glucose, mannose, raffinose, ribose, sucrose and xylose, producing mainly H2 and acetate. In addition, the isolate was able to grow with CO as the sole carbon and energy source. CO oxidation was coupled to H2 and CO2 formation. The G+C content of the genomic DNA was 54.6 mol%. Based on 16S rRNA gene sequence analysis, this bacterium is most closely related to Moorella glycerini (97 % sequence identity). Based on the physiological features and phylogenetic analysis, it is proposed that strain E3-O(T) should be classified in the genus Moorella as a representative of a novel species, Moorella stamsii. The type strain of Moorella stamsii is E3-O(T) ( = DSM 26271(T) = CGMCC 1.5181(T)).

Strategies to suppress hydrogen-consuming microorganisms affect macro and micro scale structure and microbiology of granular sludge.

Treatment of anaerobic granules with heat and two chemical treatments, contacting with 2-bromoethanesulfonate (BES) and with BES + Chloroform, were applied to suppress hydrogen-consuming microorganisms. Three mesophilic expanded granular sludge bed (EGSB) reactors-R(Heat), R(BES), and R(BES + Chlo)--were inoculated with the treated sludges and fed with synthetic sugar-based wastewater (5 g(COD) L(-1), HRT 20-12 h). Morphological integrity of granules and bacterial communities were assessed by quantitative image analysis and 16S rRNA gene based techniques, respectively. Hydrogen production in R(Heat) was under 300 mL H(2) L(-1) day(-1), with a transient peak of 1,000 mL H(2) L(-1) day(-1) after decreasing HRT. In R(BES + Chlo) hydrogen production rate did not exceed 300 mL H(2) L(-1) day(-1) and there was granule fragmentation, release of free filaments from aggregates, and decrease of granule density. In R(BES), there was an initial period with unstable hydrogen production, but a pulse of BES triggered its production rate to 700 ± 200 mL H(2) L(-1) day(-1). This strategy did not affect granules structure significantly. Bacteria branching within Clostridiaceae and Ruminococcaceae were present in this sludge. This work demonstrates that, methods applied to suppress H(2)-consuming microorganisms can cause changes in the macro- and microstructure of granular sludge, which can be incompatible with the operation of high-rate reactors.

Zeolite addition to improve biohydrogen production from dark fermentation of C5/C6-sugars and Sargassum sp. biomass.

Thermophilic biohydrogen production by dark fermentation from a mixture (1:1) of C5 (arabinose) and C6 (glucose) sugars, present in lignocellulosic hydrolysates, and from Sargassum sp. biomass, is studied in this work in batch assays and also in a continuous reactor experiment. Pursuing the interest of studying interactions between inorganic materials (adsorbents, conductive and others) and anaerobic bacteria, the biological processes were amended with variable amounts of a zeolite type-13X in the range of zeolite/inoculum (in VS) ratios (Z/I) of 0.065-0.26 g g-1. In the batch assays, the presence of the zeolite was beneficial to increase the hydrogen titer by 15-21% with C5 and C6-sugars as compared to the control, and an increase of 27% was observed in the batch fermentation of Sargassum sp. Hydrogen yields also increased by 10-26% with sugars in the presence of the zeolite. The rate of hydrogen production increased linearly with the Z/I ratios in the experiments with C5 and C6-sugars. In the batch assay with Sargassum sp., there was an optimum value of Z/I of 0.13 g g-1 where the H2 production rate observed was the highest, although all values were in a narrow range between 3.21 and 4.19 mmol L-1 day-1. The positive effect of the zeolite was also observed in a continuous high-rate reactor fed with C5 and C6-sugars. The increase of the organic loading rate (OLR) from 8.8 to 17.6 kg m-3 day-1 of COD led to lower hydrogen production rates but, upon zeolite addition (0.26 g g-1 VS inoculum), the hydrogen production increased significantly from 143 to 413 mL L-1 day-1. Interestingly, the presence of zeolite in the continuous operation had a remarkable impact in the microbial community and in the profile of fermentation products. The effect of zeolite could be related to several properties, including the porous structure and the associated surface area available for bacterial adhesion, potential release of trace elements, ion-exchanger capacity or ability to adsorb different compounds (i.e. protons). The observations opens novel perspectives and will stimulate further research not only in biohydrogen production, but broadly in the field of interactions between bacteria and inorganic materials.

Microbiologically influenced corrosion mechanism of 304L stainless steel in treated urban wastewater and protective effect of silane-TiO2 coating.

Microbiologically influenced corrosion (MIC) of bare and silane-TiO2 sol-gel coated stainless steel (SS) was studied in treated urban wastewater (TUWW). Combining the electrochemical impedance spectroscopy (EIS) and the scanning vibrating electrode technique (SVET) showed that SS surface colonization occurs, at earlier stages, by iron-oxidizing bacteria (IOB), and later by sulphate-reducing bacteria (SRB). The SVET results showed that chemical corrosion process and bacterial respiration led to the depletion of dissolved oxygen, creating a differential aeration cell and thus a localized corrosion phenomenon. Scanning electron microscopy (SEM) and energy dispersive spectroscopy (EDS) showed that the growth of a bacterial biofilm on 304L SS was a dynamic process, stimulating the localized oxidation of SS. To improve corrosion protection, a silane-TiO2 sol-gel coating for SS is proposed. SEM showed that the coating reduced bacterial adhesion and EIS study demonstrated that the coating improved the barrier properties and corrosion resistance of 304L SS in TUWW over a short period of immersion.

Advanced monitoring of high-rate anaerobic reactors through quantitative image analysis of granular sludge and multivariate statistical analysis.

Four organic loading disturbances were performed in lab-scale EGSB reactors fed with ethanol. In load disturbance 1 (LD1) and 2 (LD2), the organic loading rate (OLR) was increased between 5 and 18.5 kg COD m(-3) day(-1), through the influent ethanol concentration increase, and the hydraulic retention time decrease from 7.8 to 2.5 h, respectively. Load disturbances 3 (LD3) and 4 (LD4) were applied by increasing the OLR to 50 kg COD m(-3) day(-1) during 3 days and 16 days, respectively. The granular sludge morphology was quantified by image analysis and was related to the reactor performance, including effluent volatile suspended solids, indicator of washout events. In general, it was observed the selective washout of filamentous forms associated to granules erosion/fragmentation and to a decrease in the specific acetoclastic activity. These phenomena induced the transitory deterioration of reactor performance in LD2, LD3, and LD4, but not in LD1. Extending the exposure time in LD4 promoted acetogenesis inhibition after 144 h. The application of Principal Components Analysis determined a latent variable that encompasses a weighted sum of performance, physiological and morphological information. This new variable was highly sensitive to reactor efficiency deterioration, enclosing variations between 27% and 268% in the first hours of disturbances. The high loadings raised by image analysis parameters, especially filaments length per aggregates area (LfA), revealed that morphological changes of granular sludge, should be considered to monitor and control load disturbances in high rate anaerobic (granular) sludge bed digesters.

Garden and food waste co-fermentation for biohydrogen and biomethane production in a two-step hyperthermophilic-mesophilic process.

Co-fermentation of garden waste (GW) and food waste (FW) was assessed in a two-stage process coupling hyperthermophilic dark-fermentation and mesophilic anaerobic digestion (AD). In the first stage, biohydrogen production from individual substrates was tested at different volatile solids (VS) concentrations, using a pure culture of Caldicellulosiruptor saccharolyticus as inoculum. FW concentrations (in VS) above 2.9 g L-1 caused a lag phase of 5 days on biohydrogen production. No lag phase was observed for GW concentrations up to 25.6 g L-1. In the co-fermentation experiments, the highest hydrogen yield (46 ±â€¯1 L kg-1) was achieved for GW:FW 90:10% (w/w). In the second stage, a biomethane yield of 682 ±â€¯14 L kg-1 was obtained using the end-products of GW:FW 90:10% co-fermentation. The energy generation predictable from co-fermentation and AD of GW:FW 90:10% is 0.5 MJ kg-1 and 24.4 MJ kg-1, respectively, which represents an interesting alternative for valorisation of wastes produced locally in communities.

Electrooxidation as the anaerobic pre-treatment of fats: oleate conversion using RuO2 and IrO2 based anodes.

Electrochemical treatment of oleate using RuO2 and IrO2 type dimensionally stable anodes in alkaline medium was performed to develop a feasible anaerobic pre-treatment of fatty effluents. The results showed that the pre-treated solutions over RuO2 were faster degraded by anaerobic consortium than the raw oleate solutions or the electrolysed solutions using IrO2. In batch experiments carried out with pre-treated solutions over RuO2 (100-500mg/L), no lag phases were observed before the methane production onset. On the other hand, raw oleate and pre-treated oleate over IrO2 had originated lag phases of 0-140 and 0-210h, respectively. This study demonstrated that it is advantageous to apply the electrochemical treatment carried out on the RuO2 type DSA in order to achieve a faster biodegradation of lipid-containing effluent and consequently to obtain a faster methane production.

Photocatalytic and combined anaerobic-photocatalytic treatment of textile dyes.

A photocatalytic process based on immobilized titanium dioxide was used to treat crude solutions of azo, anthraquinone and phthalocyanine textile dyes. In addition, the process was applied to the treat autoxidized chemically reduced azo dyes, i.e. representatives of recalcitrant dye residues after biological sequential anaerobic-aerobic treatment. Photocatalysis was able to remove more than 90% color from crude as well as autoxidized chemically reduced dye solutions. UV-absorbance and COD were also removed but to a lower extent (50% in average). The end products of photocatalytic treatment were not toxic toward methanogenic bacteria. The results demonstrate that photocatalysis can be used as a pre- or post-treatment method to biological anaerobic treatment of dye-containing textile wastewater.

Influence of composition on the biomethanation potential of restaurant waste at mesophilic temperatures.

A synthetic waste was used to study the effect of waste composition on anaerobic degradation of restaurant waste. It was made by blending melted pork lard, white cabbage, chicken breast, and potato flakes, to simulate lipids, cellulose, protein, and carbohydrates, respectively. Four blends of the four constituents with an excess of each component were assayed and compared with a fifth blend containing an equal amount of chemical oxygen demand (COD) of each of the four components. The methane production and the time course of soluble COD and volatile fatty acids were assessed in batch assays. A high reduction of volatile solids (between 94% and 99.6%) was obtained in all the assays. The methane yield was between 0.40 m(3) CH(4)/kg VS(initial) (excess of carbohydrates) and 0.49 m(3) CH(4)/kg VS(initial) (excess of lipids). The degradation of the lipid-rich assays differed from the others. Fifty percent of the biochemical methane potential was obtained after 3-6 days for all of the assays, except for the one with excess of lipids which achieved 50% methanation only after 14.7 days of incubation. In the assay with excess of lipids, a considerable fraction of COD remained in the liquid phase, suggesting an inhibition of the methanogenic process that was likely due to the accumulation of long chain fatty acids. The hydrolysis rate constants, assuming first order kinetics, over the first 6 days were between 0.12d(-1) (excess of lipids) and 0.32 d(-1) (excess of carbohydrates). The results indicate that anaerobic digestion facilities with large variations in lipid input could have significant changes in process performance that merit further examination.

Anaerobic microbial LCFA degradation in bioreactors.

This paper reviews recent results obtained on long-chain fatty acids (LCFA) anaerobic degradation. Two LCFA were used as model substrates: oleate, a mono-unsaturated LCFA, and palmitate, a saturated LCFA, both abundant in LCFA-rich wastewaters. 16S rRNA gene analysis of sludge samples submitted to continuous oleate- and palmitate-feeding followed by batch degradation of the accumulated LCFA demonstrated that bacterial communities were dominated by members of the Clostridiaceae and Syntrophomonadaceae families. Archaeal populations were mainly comprised of hydrogen-consuming microorganisms belonging to the genus Methanobacterium, and acetate-utilizers from the genera Methanosaeta and Methanosarcina. Enrichment cultures growing on oleate and palmitate, in the absence or presence of sulfate, gave more insight into the major players involved in the degradation of unsaturated and saturated LCFA. Syntrophomonas-related species were identified as predominant microorganisms in all the enrichment cultures. Microorganisms clustering within the family Syntrophobacteraceae were identified in the methanogenic and sulfate-reducing enrichments growing on palmitate. Distinct bacterial consortia were developed in oleate and palmitate enrichments, and observed differences might be related to the different degrees of saturation of these two LCFA. A new obligately syntrophic bacterium, Syntrophomonas zehnderi, was isolated from an oleate-degrading culture and its presence in oleate-degrading sludges detected by 16S rRNA gene cloning and sequencing.

Activated sludge process monitoring through in situ near-infrared spectral analysis.

The application of near infrared (NIR) spectroscopy for industrial process monitoring is achieving increasing importance over the last twenty years. In fact, the real time monitoring capacity of NIR spectroscopy is a very important feature for process monitoring, prediction and control as it allows a fast evaluation of the state of the process. However, the application of NIR spectroscopy in wastewater treatment processes is still to be explored. Although some applications of the technique for wastewater monitoring have been reported in the literature, there is still a need for more investigation related with applications, limitations and advantages of the technique when compared with other methods. An activated sludge reactor for aerobic treatment of a complex medium was monitored in situ with a NIR transflectance probe and traditional chemical parameters analysed off-line. NIR spectrophotometric data measured at the feed, reactor and settler were coupled to principal component analysis (PCA) to infer about the ability of this monitoring system to detect changes in the feed influent. The analysis of the score plots resulting from PCA permitted to identify the moments at which the perturbations occurred and to follow the consequent instability induced in the reactor till the day where the system is recuperated. The promising results obtained, suggest the interest in more detailed studies on the feasibility of NIR spectroscopy as an alternative method for monitoring and control of wastewater treatment processes.

Protein from Hevea brasiliensis "Hev b 13" latex attenuates systemic inflammatory response and lung lesions in rats with sepsis.

Sepsis induces a severe systemic inflammatory response that may result in multiple organ dysfunction and death. Studies using a protein derived from natural Hevea brasiliensis (rubber tree) latex, denominated Hev b 13, have demonstrated important anti-inflammatory effects, but no data have been published regarding its effects on sepsis. The aim of this study was to investigate the effects of Hev b 13 on the inflammatory response and lung lesions of septal rats. Male Wistar rats were submitted to cecal ligation and puncture (CLP), randomized into groups and treated with subcutaneously administered doses of 0.5/2.0/3.0 mg/Kg of Hev b 13. Next, animals were subdivided into three different points in time (1, 6 and 24 hours after treatments) for collection of blood samples and euthanasia accompanied by organ removal. Total and differential leukocyte counts, cytokine dosage and histological assessment were analyzed. Treatment with Hev b 13 resulted in a significant decline in total and differential leukocytes as well as suppression of TNF-α and IL-6 production, associated with the increase in IL-10 and IL-4 in plasma and lung tissue. Moreover, it reduced morphological and pathological changes found in the lungs, including neutrophil infiltration, edema and alveolar thickening. The present study concluded that Hev b 13 exerts anti-inflammatory effects and attenuates lung lesions in septal rats, showing potential for clinical application.

Potential anti-cancer and anti-Candida activity of Zn-derived foams.

Zinc (Zn)-derived foams have been prepared from an alkaline electrolyte solution by galvanostatic electrodeposition under different conditions. A detailed physico-chemical characterization was performed by Raman spectroscopy, X-ray diffraction (XRD) and scanning electron microscopy (SEM). A pioneer application of these foams in medical implant-related applications was investigated. The in vitro behaviour of these Zn-derived foams in simulated physiological conditions was studied. The results revealed that the presence of zinc oxide was important enough to change the in vitro behaviour of these materials. The potential of these Zn-derived foams in inhibiting bone cancer cell proliferation - osteoscarcoma cells - and important pathogenic fungi responsible for implant-related infections -Candida albicans- was examined. Furthermore, the foams were evaluated for cytocompatibility with normal human osteoblasts. The results obtained allowed us to conclude that Zn-derived foams have an interesting potential for anti-cancer and anti-Candida activity, targeted for bone-related implant applications, suggesting that this novel material may have potential for further clinical studies.

Quantitative image analysis as a diagnostic tool for identifying structural changes during a revival process of anaerobic granular sludge.

Due to unspecified operational problems, the specific acetoclastic activity (SAA) of the anaerobic granular sludge present in an industrial UASB reactor was considerably damaged (from 250 to less than 10mL CH(4)@STP/gVSS.d), significantly reducing the biogas production of that industrial unit. The hydrogenotrophic methanogenic activity exhibited a value of 600mL CH4@STP/gVSS.d, the settling velocity was 31.4+/-9.8m/h, the average equivalent diameter was 0.92+/-0.43mm, and about 70% of the VSS were structured in aggregates larger than 1mm. In order to study the recovery of the SAA, this sludge was collected and inoculated in a lab-scale expanded granular sludge blanket (EGSB) reactor. Ethanol was fed as the sole carbon source during a trial period of 106 days. Process monitoring included COD removal efficiency, methane production, and periodic determination of the specific methanogenic activity in the presence of acetate, propionate, butyrate, ethanol and H(2)/CO(2). Quantitative image analysis allowed for information to be obtained on granular fragmentation/erosion and filaments release. During the first operational period, biogas production was mainly due to the hydrogenotrophic activity. However, after 40 days, the SAA steadily increased achieving a maximum value of 183+/-13mL CH4@STP/gVSS.d. The onset of SAA recovery, granules breakdown and filaments release to the bulk occurred simultaneously. Further increase in SAA was accompanied by granular growth. In the last 25 days of operation, the size distribution was stable with more than 80% of projected area of aggregates corresponding to granules larger than 1mm (equivalent diameter). Confocal images from FISH hybridized sections of the granules showed that after SAA recovery, the granules developed an organized structure where an acidogenic/acetogenic external layer was apparent. Granular fragmentation and increase of filaments in the bulk, simultaneously with the increase in the acetoclastic activity are described for the first time and might represent a structural response of granular sludge to promote the optimal substrate uptake at minimal diffusion limitations.

Anaerobic co-digestion of coffee waste and sewage sludge.

The feasibility of the anaerobic co-digestion of coffee solid waste and sewage sludge was assessed. Five different solid wastes with different chemical properties were studied in mesophilic batch assays, providing basic data on the methane production, reduction of total and volatile solids and hydrolysis rate constant. Most of the wastes had a methane yield of 0.24-0.28 m3 CH4(STP)/kg VS(initial) and 76-89% of the theoretical methane yield was achieved. Reduction of 50-73% in total solids and 75-80% in volatile solids were obtained and the hydrolysis rate constants were in the range of 0.035-0.063 d(-1). One of the solid wastes, composed of 100% barley, achieved a methane yield of 0.02 m3 CH4(STP)/kg VS(initial), reductions of 31% in total solids, 40% in volatile solids and achieved only 11% of the theoretical methane yield. However, this waste presented the highest hydrolysis rate constant. Considering all the wastes, an inverse linear correlation was obtained between methane yield and the hydrolysis rate constant, suggesting that hydrolysis was not the limiting factor in the anaerobic biodegradability of this type of waste.

Knowledge-based fuzzy system for diagnosis and control of an integrated biological wastewater treatment process.

A supervisory expert system based on fuzzy logic rules was developed for diagnosis and control of a laboratory- scale plant comprising anaerobic digestion and anoxic/aerobic modules for combined high rate biological N and C removal. The design and implementation of a computational environment in LabVIEW for data acquisition, plant operation and distributed equipment control is described. A step increase in ammonia concentration from 20 to 60 mg N/L was applied during a trial period of 73 h. Recycle flow rate from the aerobic to the anoxic module and bypass flow rate from the influent directly to the anoxic reactor were the output variables of the fuzzy system. They were automatically changed (from 34 to 111 L/day and from 8 to 13 L/day, respectively), when new plant conditions were recognised by the expert system. Denitrification efficiency higher than 85% was achieved 30 h after the disturbance and 15 h after the system response at an HRT as low as 1.5 h. Nitrification efficiency gradually increased from 12 to 50% at an HRT of 3 h. The system proved to react properly in order to set adequate operating conditions that led to timely and efficient recovery of N and C removal rates.

Effect of electrochemical treatment of oleic acid on anaerobic digestion.

Electrochemical pre-treatment of oleate on anaerobic digestion was investigated using two anaerobic filters (AF and AFm) working, respectively, with increasing concentrations of electrochemically pre-treated oleate and oleate. Influents containing 25% and 50% of chemical oxygen demand (COD) of these substrates did not cause relevant alterations in either digester's performance. Comparatively, the pre-treated oleate unit (AF) showed a higher gas production but a lower COD removal than AFm (2.0 vs 1.5 m(3) CH4 m(-3) d(-1) 76-85 vs 91-93% COD removal). The raised AFm COD removal together with the larger proportion of VSS in its effluent than in AF (5.69 vs 0.26 kg m(-3)) indicate that the biomass washout can be a consequence of the encapsulated solids by lipidic compounds that were not fairly degraded. Further increases of oleate in AFm feed (75 and 100% COD) led to decreases of biogas production to half and COD removal from values higher than 90% to 83-75%. The simultaneous increase of effluent VSS concentrations to 10-12 kg m(3) emphasizes the oleate negative effect. The use of pretreated oleate as the only carbon source did not influence negatively the AF behaviour in terms of methane production. COD removal was maintained at values higher than 80% and the effluent VSS at low concentrations (0.15 kg m(-3)). The significantly higher methane yield achieved by AF than AFm indicates a higher accumulation of oleate than pre-treated oleate. Therefore, electrochemical process stands for a suitable pre-treatment for lipid rich effluents, allowing the application of higher organic loads keeping a higher methane yield.