Formation of Recalcitrant Compounds during Anaerobic Digestion of Thermally Pre-Treated Sludge: A Critical Macromolecular and Structural Study.

Thermal hydrolysis, when used as pre-treatment, enhances the anaerobic digestion of sewage sludge; moreover, due to the high temperature normally applied, undesirable recalcitrant compounds via Maillard reactions may also be formed. However, although the appearance of these recalcitrant compounds is widely reported, more information on the formation, structure, and fate of these compounds is still needed. This study was focused on understanding the amount and whereabouts of such compounds during the anaerobic digestion process with thermal pre-treatment in soluble and total phase and advance in its structural identification by analyzing their infrared (IR) spectra. It was found that, even with the improved methane production and COD degradation, at 165 °C for 30 min, humic-like compounds are formed which could not be degraded at the anaerobic digestion step. These compounds account for 25% of the original sludge. Infrared spectroscopy proved to be a powerful technique, permitting their differentiation from the natural humic-like compounds. This research provides new information about the structure of melanoidins at every stage of the thermal hydrolysis pre-treatment and how they contribute to the dissolved organic nitrogen.

On-Line Control of Feast/Famine Cycles to Improve PHB Accumulation during Cultivation of Mixed Microbial Cultures in Sequential Batch Reactors.

Production of polyhydroxyalkanoates (PHA) has generated great interest as building blocks for bioplastic production. Their production using mixed microbial cultures represents an interesting alternative, since it enables the use of organic wastes as a carbon source. Feast/famine strategy is a common way to promote selection of microorganisms with PHA accumulation capacity. However, when using waste sources, changes in substrate concentration are expected, that may affect performance and efficiency of the process. This study showed how the dissolved oxygen level can be used for online control of the cycle time, ensuring that the desired feast/famine ratio is effectively applied. An operation strategy is presented and validated, using sequential batch reactors fed with acetate as the carbon source. Production of polyhydroxybutyrate (PHB) was studied, which is the expected type of PHA to be synthetized when using acetate as substrate. Two reactors were operated by applying the proposed control strategy, to provide F/F ratios of 0.2 and 0.6, respectively. A third reactor was operated with a fixed cycle time, for comparison purposes. Results showed that the reactor that operated at an F/F ratio of 0.6 promoted higher biomass productivity and PHB content, as a result of a better use of available time, preventing unnecessary long famine times. The application of the tested strategy is a simple a reliable way to promote a better performance of feast/famine-based bioreactors involving mixed microbial cultures for PHB production.

Maximizing energy recovery from wastewater via bioflocculation-enhanced primary treatment: a pilot scale study.

Anaerobic digestion of municipal sewage sludge is widely used for harvesting energy from wastewater organic content. The more organic carbon we can redirect into the primary sludge, the less energy is needed for aeration in secondary treatment and the more methane is produced in anaerobic digesters. Bioflocculation has been proposed as a promising separation technology to maximize carbon capture in primary sludge. Thus far, only limited data on bioflocculation are available under real conditions, i.e. from pilot-scale reactors treating raw sewage. Moreover, no study has discussed yet the influence of bioflocculation on denitrification potential of sewage. Therefore, we performed bioflocculation of raw sewage in high-rate contact stabilization process in pilot-scale to investigate maximal primary treatment efficiency. During 100 days of operation at sludge retention time of only 2 days, the average removal efficiencies of chemical oxygen demand (COD), suspended solids and total phosphorus were 75%, 87% and 51%, respectively, using no chemicals for precipitation. Up to 76% of incoming COD was captured in primary sludge and 46% for subsequent anaerobic digestion, where energy recovery potential achieved 0.33-0.37 g COD as CH4 per g COD of influent. This study showed in real conditions that this newly adapted separation process has significant benefits over chemically enhanced primary treatment, enabling sewage treatment process to overcome energy self-sufficiency.

Micro-Oxygenation in Upflow Anaerobic Sludge Bed (UASB) Reactors Using a Silicon Membrane for Sulfide Oxidation.

Sulfide produced by sulphate-reducing bacteria in anaerobic reactors can seriously affect biogas quality. Microaeration has become a reliable way to remove sulfide, by promoting its oxidation. However, limited research is available regarding its application in upflow anaerobic sludge bed (UASB) reactors. In this research, silicon membranes were studied as a mechanism to dose oxygen in USAB reactors. Two configurations were tested: the membrane placed inside the reactor or in an external module. Our results show that the external membrane proved to be a more practical alternative, providing conditions for sulfide oxidation. This led to a reduction in its concentration in the liquid effluent and biogas. External membrane configuration achieved a sulfide conversion rate of 2.4 g-S m2 d-1. Since the membrane was not sulfide-selective, methane losses were observed (about 9%). In addition, excessive oxygen consumption was observed, compared to the stoichiometric requirement. As is the case for many membrane-based systems, membrane area is a key factor determining the correct operation of the system.

Making sense of parameter estimation and model simulation in bioprocesses.

Most articles that report fitted parameters for kinetic models do not include meaningful statistical information. This study demonstrates the importance of reporting a complete statistical analysis and shows a methodology to perform it, using functionalities implemented in computational tools. As an example, alginate production is studied in a batch stirred-tank fermenter and modeled using the kinetic model proposed by Klimek and Ollis (1980). The model parameters and their 95% confidence intervals are estimated by nonlinear regression. The significance of the parameters value is checked using a hypothesis test. The uncertainty of the parameters is propagated to the output model variables through prediction intervals, showing that the kinetic model of Klimek and Ollis (1980) can simulate with high certainty the dynamic of the alginate production process. Finally, the results obtained in other studies are compared to show how the lack of statistical analysis can hold back a deeper understanding about bioprocesses.

Application of microbe-induced carbonate precipitation for copper removal from copper-enriched waters: Challenges to future industrial application.

Copper contamination in watercourses is a recent issue in countries where mining operations are prevalent. In this study, the application of copper precipitation through microbe-induced carbonate precipitation (MICP) was analyzed using urea hydrolysis by bacteria to evaluate precipitated copper carbonates. This article demonstrates the application of a copper precipitation assay involving Sporosarcina pasteurii (in 0.5 mM Cu2+ and 333 mM urea) and analyzes the resultant low removal (10%). The analysis indicates that the low removal was a consequence of Cu2+ complexation with the ammonia resulting from the hydrolysis of urea. However, the results indicate that there should be a positive correlation between the initial urea concentration and the bacterial tolerance to copper. This identifies a challenge in the industrial application of the process, wherein a minimum consumption of urea represents an economic advantage. Therefore, it is necessary to design a sequential process that decouples bacterial growth and copper precipitation, thereby decreasing the urea requirement.

Effect of Operational Conditions on the Behaviour and Associated Costs of Mixed Microbial Cultures for PHA Production.

Massive production and disposal of petrochemical derived plastics represent relevant environmental problems. Polyhydroxyalkanoates (PHA) are a renewable alternative that can even be produced from wastes. The production of PHA from acetate using mixed microbial cultures was studied. The effect of two key operational conditions was evaluated, i.e., substrate concentration and cycle length. The effects of these factors on several responses were studied using a surface response methodology. Several reactors were operated under selected conditions for at least 10 solids retention times to ensure stable operation. Results show that conditions providing higher PHA content involve lower biomass productivities. This has a great impact on biomass production costs. Results suggest then that PHA content alone may not be a reasonable criterion for determining optimal conditions for PHB production. If production costs need to be reduced, conditions that provide a lower PHA content in the selection reactor, but a higher biomass productivity may be of interest.

The accumulation of volatile fatty acids and phenols through a pH-controlled fermentation of olive mill solid waste.

This work aims to compare the use of olive mill solid waste as substrate in pH-controlled fermentation at acid (pH = 5), neutral (uncontrolled, pH ≈ 7) and alkaline (pH = 9) operating pH levels. The results obtained in this study indicate that operating pH strongly affected the anaerobic microorganisms and, hence, different target compounds could be obtained by adjusting the operating pH. Fermentation at neutral pH resulted in the conversion of 93.5% of the fed chemical oxygen demand to methane. However, fermentations at pH 5 and 9 resulted in the inhibition of the methanogenic activity. At pH 9, volatile fatty acids reached a maximum concentration of 3.69 g O2/L, where acetic acid represented up to 79.3% of the total volatile fatty acids. Unlike volatile fatty acid production, an optimal operation of fermentation at pH 5 could allow the recovery of phenols such as vanillin.

A membrane aerated biofilm reactor for sulfide control from anaerobically treated wastewater.

A upflow anaerobic sludge blanket reactor was operated combined to a membrane aerated biofilm reactor for sulfate removal and for elemental sulfur reclamation. A commercial silicon tube was used as an oxygen delivery diffuser. The process achieved high rates of sulfide removal from the liquid phase (90%). The hydrogen sulfide removal was influenced by the pH value and at pH value of 7.5, 98% of the H2S was removed. The elemental sulfur was observed inside the membrane, with content in the biomass of 21%. Through the massive sequencing of the samples, the microbial community diversity and the stratification of biomass inside the silicon tube was demonstrated, confirming the presence of sulfide-oxidizing bacteria on the membrane wall. The most important genera found related to the sulfur cycle were Sulfuricurvum, Geovibrio, Flexispira and Sulforospirillum.

A need for a standardization in anaerobic digestion experiments? Let's get some insight from meta-analysis and multivariate analysis.

An important variability in the experimental results in anaerobic digestion lab test has been reported. This study presents a meta-analysis coupled with multivariate analysis aiming to assess the impact of this experimental variability in batch and continuous operation at mesophilic and thermophilic anaerobic digestion of waste activated sludge. An analysis of variance showed that there was no significant difference between mesophilic and thermophilic conditions in both continuous and batch conditions. Concerning the operation mode, the values of methane yield were significantly higher in batch experiment than in continuous reactors. According to the PCA, for both cases, the methane yield is positive correlated to the temperature rises. Interestingly, in the batch experiments, the higher the volatile solids in the substrate was, the lowest was the methane production, which is correlated to experimental flaws when setting up those tests. In continuous mode, unlike the batch test, the methane yield is strongly (positively) correlated to the organic content of the substrate. Experimental standardization, above all, in batch conditions are urgently necessary or move to continuous experiments for reporting results. The modeling can also be a source of disturbance in batch test.

Use of activated sludge biomass as an agent for advanced primary separation.

Conventional primary settling is a physical process of solid-liquid separation, normally presenting low removal efficiencies. Improvement of this separation process would result in energetic advantages: lower aeration requirements and higher biogas production form primary and secondary sludges. Secondary sludge has been proposed as a potential agent promoting an increase in primary separation efficiency. Few processes have been proposed, based on the cultivation of sludge under special conditions. However, one can speculate that regular sludge may have a similar effect. The aim of this research was to study that possibility. Sludges from different activated sludge reactors were tested. Results showed that COD removals were up to 55%, 2 times higher than that for simple settling. Under that condition, COD balances showed that aeration requirements would reduce 40%, and biogas production from primary and secondary sludges would increase 50%. It is inferred then that the application of activated sludge as an external agent represents an interesting alternative that have the potential to significantly improve energetic efficiency of sewage treatment plants.

Screening of biomethane production potential from dominant microalgae.

The use of microalgae for biomethane production has been considerably increasing during the recent years. In this study, four dominant species belonging to the genera Scenedesmus, Chlorella, Dunaliella and Nostoc were selected. The influence of different genera with several morphological, structural and physicochemical characteristics on methane production was assessed in biochemical methane potential (BMP) tests. The ultimate methane yield values were 332 ± 24, 211 ± 2, 63 ± 17 and 28 ± 10 mL CH4/g VSadded for Scenedesmus obliquus, Chlorella sorokiniana, Dunaliella salina and Nostoc sp., respectively. The highest methane production was achieved by microalga species that had no complex cell wall or wall basically composed by proteins and simple sugars such as in S. obliquus, whereas lower methane yields were found for D. salina and Nostoc sp., due to the salinity effects and cell wall composition in terms of complex polysaccharide and glycolipid layers, respectively. Kinetic constant values obtained in the BMP tests ranged between 1.00 ± 0.08 and 0.097 ± 0.005 days(-1) for D. salina and S. obliquus, respectively.

Batch anaerobic co-digestion of waste activated sludge and microalgae (Chlorella sorokiniana) at mesophilic temperature.

The microalgae Chlorella sorokiniana are used as co-substrate for waste activated sludge (WAS) anaerobic digestion. The specific objective of this research was to evaluate the feasibility of improving methane production from anaerobic digestion of WAS in co-digestion with this microalga, based on an optimized mixture percentage. Thus, the anaerobic co-digestion of both substrates aims to overcome the drawbacks of the anaerobic digestion of single WAS, simultaneously improving its management. Different co-digestion mixtures (0% WAS-100% microalgae; 25% WAS-75% microalgae; 50% WAS-50% microalgae; 75% WAS-25% microalgae; 100% WAS-0% microalgae) were studied. The highest methane yield (442 mL CH4/g VS) was obtained for the mixture with 75% WAS and 25% microalgae. This value was 22% and 39% higher than that obtained in the anaerobic digestion of the sole substrates WAS and microalgae, respectively, as well as 16% and 25% higher than those obtained for the co-digestion mixtures with 25% WAS and 75% microalgae and 50% WAS and 50% microalgae, respectively. The kinetic constant of the process increased 42%, 42% and 12%, respectively, for the mixtures with 25%, 50% and 75% of WAS compared to the substrate without WAS. Anaerobic digestion of WAS, together with C. sorokiniana, has been clearly improved by ensuring its viability, suitability and efficiency.

Use of cellulolytic marine bacteria for enzymatic pretreatment in microalgal biogas production.

In this study, we designed and evaluated a microalgal pretreatment method using cellulolytic bacteria that naturally degrades microalgae in their native habitat. Bacterial strains were isolated from each of two mollusk species in a medium containing 1% carboxymethyl cellulose agar. We selected nine bacterial strains that had endoglucanase activity: five strains from Mytilus chilensis, a Chilean mussel, and four strains from Mesodesma donacium, a clam found in the Southern Pacific. These strains were identified phylogenetically as belonging to the genera Aeromonas, Pseudomonas, Chryseobacterium, and Raoultella. The cellulase-producing capacities of these strains were characterized, and the degradation of cell walls in Botryococcus braunii and Nannochloropsis gaditana was tested with "whole-cell" cellulolytic experiments. Aeromonas bivalvium MA2, Raoultella ornithinolytica MA5, and Aeromonas salmonicida MC25 degraded B. braunii, and R. ornithinolytica MC3 and MA5 degraded N. gaditana. In addition, N. gaditana was pretreated with R. ornithinolytica strains MC3 and MA5 and was then subjected to an anaerobic digestion process, which increased the yield of methane by 140.32% and 158.68%, respectively, over that from nonpretreated microalgae. Therefore, a "whole-cell" cellulolytic pretreatment can increase the performance and efficiency of biogas production.

Enzymatic biodiesel production kinetics using co-solvent and an anhydrous medium: a strategy to improve lipase performance in a semi-continuous reactor.

Enzymatic biodiesel production kinetics under previously optimized conditions were investigated. Waste frying oil (WFO) was used as the raw material, Novozym 435 as catalyst, methanol as acyl acceptor and tert-butanol as co-solvent. To investigate pure transesterification kinetics improving product properties, 3Å molecular sieves were incorporated into the reaction to provide an anhydrous medium avoiding the side reactions of hydrolysis and esterification. The effects of either WFO or methanol on the reaction rate were analyzed separately. The reaction was described by a Ping Pong mechanism and competitive inhibition by methanol. The results obtained in the kinetics study were applied in the operation of a semi-continuous reactor for biodiesel production. The operational conditions of each reaction cycle were: methanol-to-oil ratio 8/1 (mol/mol), 15% (wt) Novozym 435, 0.75% (v/v) of tert-butanol, 44.5°C, 200 rpm and 4h of reaction time. The enzymes were successively reused by remaining in the reactor during all the cycles. Under these conditions, biodiesel production yields higher than 80% over 7 reaction cycles were observed. Both the kinetics study and the reactor operation showed that Novozym 435 was not inhibited at high methanol concentrations and that the kinetics of the proposed enzymatic process could be comparable to the conventional chemical process.

Behavior of the anaerobic treatment of tannery wastewater at different initial pH values and sulfate concentrations.

The influence of initial pH and sulfate concentration on the anaerobic digestion of tannery wastewater was studied using the specific methanogenic activity (SMA) test. The SMA tests were performed with a chemical oxygen demand (COD) concentration of 6.5 g/L and SO(-2)4 concentrations of 1.95, 3.90, 6.50, 8.45 and 10.4 g/L. The effect of pH was studied at pH 6.8; 6.4 and 6.1. It was found that methane production took place under all conditions, but the use of SO(-2)4 concentrations higher than 8.45 g/L and initial pH levels lower than 6.4 caused a decrease of up to 73% of methane production. The influence of pH was significant only for SO(-2 4 concentrations lower than 8.45 g/L, with up to 50% inhibition. Specific methanogenic activity for COD/SO(-2)4 ratios lower than 0.77 depends basically on this ratio and not on the initial pH value under the conditions of the study.

Potentials of anaerobic membrane bioreactors to overcome treatment limitations induced by industrial wastewaters.

This review presents a comprehensive summary on applications of anaerobic membrane bioreactor (AnMBR) technology for industrial wastewaters in view of different aspects including treatability and filterability. AnMBRs present an attractive option for the treatment of industrial wastewaters at extreme conditions, such as high salinity, high temperature, high suspended solids concentrations, and toxicity that hamper granulation and retention of biomass or reduce the biological activity. So far, most of the research has been conducted at laboratory scale; however, also a number of full-scale AnMBR systems is currently being operated worldwide. Membrane fouling, a multivariable process, is still a research quest that requires further investigation. In fact, membrane fouling and flux decline present the most important reasons that hamper the wide-spread application of full-scale reactors. This paper addresses a detailed assessment and discussion on treatability and filterability of industrial wastewaters in both lab- and full-scale AnMBR applications, the encountered problems and future opportunities.

Biofilm formation for organic matter and sulphate removal in gas-lift reactors

A start-up strategy was presented and evaluated to obtain a well-established biofilm in a gas lift-reactor capable both for the removal of organic matter and sulphate. Pumice stone was used as material support. The influence of shear forces, given by the biogas recirculation, the effect of the COD/SO4-2 ratio and the OLRs increase were evaluated on the reactor performance. From the first stages, cell colonization was observed along with the presence of extracellular polymeric substances. The COD and sulphate removal was over 70 percent, for all conditions. The increase of gas flow did not have an adverse effect on biofilm development even though there was some detachment. Specific methanogenic activity of the biofilm increased along the experiments. Operational parameters as alkalinity and alkalinity ratio were within the recommended values for the operation with sulphate-rich wastewater. For gas-lift reactors operation it becomes fundamental to have a suitable start-up strategy that takes into account the initial biofilm development from a non-acclimatized biomass.

Simultaneous effects of pH and substrate concentration on hydrogen production by acidogenic fermentation / Efectos simultáneos del pH y la concentración de sustrato en la producción de hidrógeno mediante fermentación acidogénica

The present research examined the effects of initial substrate concentration and pH on the yield and productivity of hydrogen production by acidogenic fermentation. Assays were carried out at three different initial pH levels (5.5, 6.5 and 7.5) and three initial substrate concentrations (3, 5 and 10 g COD/L). Glucose was used as carbon source and the experiments were conducted at 37°C in batch tests, after a thermal pretreatment to eliminate methanogenic microorganisms. Conversions of glucose into hydrogen were between 16.75 and 27.25 percent of theoretical maximum, and high values of hydrogen productivity were obtained. An optimum value for the yield of glucose between initial pH of 6.3 and 3.7 g COD/L and productivity of the 5.95 H2/gVSS h and initial pH of 6.7 and 10 g COD/L were obtained from the response surface.

Effects of the acidogenic biomass on the performance of an anaerobic membrane bioreactor for wastewater treatment.

Continuous flow experiments were performed to study the effects of acidogenic biomass development, induced by feeding with non-acidified substrate, on the operation and performance of an anaerobic membrane bioreactor (AnMBR). The AnMBR was operated at cross-flow velocities up to 1.5m/s and fed with a gelatine-starch-ethanol mixture. A significant fraction of acidogenic biomass developed during reactor operation, which fully determined the sludge rheology, and influenced the particle size distribution. As a result, flux levels of only 6.5l/m(2)h were achieved, at a liquid superficial velocity of 1.5m/s. Even though the soluble microbial products levels in the AMBR were as high as 14g COD/l, the observed hydraulic flux was not limited by irreversible pore fouling, but by reversible cake layer formation. Propionate oxidation was the limiting step for the applied organic loading rate. The assessed specific methanogenic activity (SMA) with propionate as substrate was, however, similar to the values found by others during thermophilic treatment of non or partially acidified substrates in granular sludge bed reactors, indicating an appropriate level of the propionate oxidation capacity.