Xylo-oligosaccharides (XOs) from xylans are gaining interest due to their wide use in food, feed and pharmaceutical industry. In this work, the enzymatic production of XOs from beechwood xylan in batch, continuous and semi-continuous enzyme membrane reactor mode was compared. The potential of an inline viscometer for real-time monitoring of the hydrolysis was assessed as well. In all modes, an initial concentration of 7% (w/w) beechwood xylan was applied at a temperature of 50 degrees C and a pH of 5.8. Initially, the effect of enzyme dosage was investigated in batch mode (0.5, 2.6, 12, 26 and 720 U of Cellic (R) CTec2.g(-1) xylan). Higher enzyme dosages led to a faster hydrolysis rate, but also to an undesirable higher production of xylose. The introduction of enzyme membrane reactors allowed production of XOs with weight average molecular weights of ca. 5-8.9 kDa combined with negligible xylose quantities (<3% in freeze dried product). The semi-continuous configuration has the edge in comparison to the full continuous configuration in terms of enzyme consumption and ease of operation. In vitro fermentations with three different gut bacteria were performed and confirmed the prebiotic properties of the produced XOs with average degree of polymerization of 48.

Rhodomonas salina is a microalgal species, belonging to the cryptophytes, and is widely used as aquaculture feed because of its high nutritional profile and phycoerythrin content. This study investigated the effect of pH on the growth, biochemical composition, and taste of R. salina when cultivated on a semi-large scale under natural light conditions. Two tubular photobioreactors (200 L) were used for the cultivation of R. salina with sunlight as the only illumination source. Two different pH setpoints were applied, 7 and 8.5. Optimal temperature and nutrient conditions were applied, according to previous research findings. The results demonstrated that the productivity of R. salina was higher at pH 7, 0.06–0.14 gdry weight L−1 day−1, compared to pH 8.5, 0.03–0.12 gdry weight L−1 day−1. It was found that protein and total fatty acid concentrations were higher in the biomass that was produced at pH 8.5, 33.7% and 12.3% of dry weight, respectively, while at pH 7, the protein content was 31.9% and the total fatty acids 8.8% of dry weight. The phycoerythrin concentration, like protein, was higher at pH 8.5, 2.7% of dry weight, compared to pH 7, 1% of dry weight. The free amino acid and nucleotide profile of R. salina was affected by the pH, resulting in increased equivalent umami concentration at pH 7. For the sensory evaluation, an expert panel on algae flavors evaluated the effect of pH on the taste of R. salina, reporting that the biomass that was produced at pH 7 had more umami flavor than the biomass that was produced at pH 8.5, which was evaluated as more bitter.

Sugar beet pulp pectin is an attractive source for the production of pectic oligosaccharides, an emerging class of potential prebiotics. The main aim of the present work was to investigate a new process allowing to produce pectic oligosaccharides in a continuous way by means of a cross flow enzyme membrane reactor while using a low-cost crude enzyme mixture (viscozyme). Preliminary experiments in batch and semi-continuous setups allowed to identify suitable enzyme concentrations and assessing filtration suitability. Then, in continuous experiments in the enzyme membrane reactor, residence time and substrate loading were further optimized. The composition of the obtained oligosaccharide mixtures was assessed at the molecular level for the most promising conditions and was shown to be dominated by condition-specific arabinans, rhamnogalacturonans, and galacturonans. A continuous and stable production was performed for 28.5 h at the optimized conditions, obtaining an average pectic oligosaccharide yield of 82.9 ± 9.9% (w/w), a volumetric productivity of 17.5 ± 2.1 g/L/h, and a specific productivity of 8.0 ± 1.0 g/g E/h. This work demonstrated for the first time the continuous and stable production of oligosaccharide mixtures from sugar beet pulp using enzyme membrane reactor technology in a setup suitable for upscaling.

Microalgae accumulate a variety of compounds. A cascading biorefinery approach aims at recovering multiple algae compounds to valorize the complete biomass and to improve the economics of the values chain. Different aspects of cascading biorefinery approaches are discussed, including (1) the impact of multiple compound recovery in a cascading approach on extraction technologies compared to single compounds approaches, (2) biorefinery concepts, (3) examples of biorefinery concepts that were evaluated on a lab scale, (4) economic aspects, and (5) future needs. Although theoretical algal biorefinery concepts have been reported frequently, publications describing experimental data supporting the feasibility of the concepts are rather poor and mostly related to small-scale experiments. There is a need for more experimental data, up-scaling, demonstration of potentially viable value chains, and production of larger quantities of algae-based products for application tests aiming at downstream market development.

The aim of this research was to valorize onion skins, an under-utilized agricultural by-product, into pectic oligosaccharides (POS), compounds with potential health benefits. To achieve high hydrolysis performance with the multi-activity enzyme Viscozyme L, an innovative approach was investigated based on a cross-flow continuous membrane enzyme bioreactor (EMR). The influence of the various process conditions (residence time, enzyme concentration, substrate concentration) was investigated on productivity and yield. The composition of the POS mixtures in terms of mono-and oligosaccharides was assessed at the molecular level. At optimized conditions, a stable POS production with 22.0 g/L/h volumetric productivity and 4.5 g/g POS/monosaccharides was achieved. Compared to previous results obtained in batch for the enzyme Viscozyme L, EMR provided a 3-5x higher volumetric productivity for the smallest POS. Moreover, it gave competitive results even when compared to batch production with a pure endo-galacturonase enzyme, demonstrating its feasibility for efficient POS production.

The primary purpose of this study was to evaluate the performance of composites, as electrode materials, made with activated carbon (AC) and SiO2 or γ-AlOOH gels as binders and compare these to traditional electrodes fabricated with AC and polyvinylidene fluoride (PVDF). Electrodes made out of AC and SiO2 or γ-AlOOH composites showed a much better wear resistance than those fabricated with AC and PVDF composites. Furthermore, cyclic voltammetry and chronopotentiometric measurements showed that the electrodes using oxides as binders had an electrochemical capacitance between 2 and 5 times higher than those made with AC and PVDF composites. This higher electrochemical capacitance is based on the fact that composites that use SiO2 and γ-AlOOH as binders exhibit a much higher specific surface area and water hydrophilicity than those containing PVDF. The addition of PVDF to the AC decreases its specific surface area by a factor of seven. In contrast, the addition of SiO2 or γ-AlOOH gels to the AC powder slightly increases the surface area. Scanning electron microscopy (SEM) images showed that the AC particles are coated with a nanoporous layer of these metal oxides.

The influence of different extraction methodologies was assessed on the composition of both neutral (arabinose, rhamnose, galactose) and acidic (galacturonic acid) pectic polysaccharides obtained from four agro-industrial residues, namely, berry pomace (BP), onion hulls (OH), pressed pumpkin (PP), and sugar beet pulp (SBP). For acidic pectic polysaccharides, the extraction efficiency was obtained as BP (nitric acid-assisted extraction, 2 h, 62.9%), PP (enzymatic-assisted extraction, 12 h, 75.0%), SBP (enzymatic-assisted extraction, 48 h, 89.8%; and nitric acid-assisted extraction, 4 h, 76.5%), and OH (sodium hexametaphosphate-assisted extraction, 0.5 h, 100%; and ammonium oxalate-assisted extraction, 0.5 h, 100%). For neutral pectic polysaccharides, the following results were achieved: BP (enzymatic-assisted extraction, 24 h, 85.9%), PP (nitric acid-assisted extraction, 6 h, 82.2%), and SBP (enzymatic assisted extraction, 48 h, 97.5%; and nitric acid-assisted extraction, 4 h, 83.2%). On the basis of the high recovery of pectic sugars, SBP and OH are interesting candidates for the further purification of pectin and production of pectin-derived products.

Micro algae are a single celled biomass with a very high potential in the biobased economy. In algal biomass cultivation, the harvesting step (i.e., separation of the dilute single cell algae from the growth medium) is a considerable part of the capital and operational cost. These processes typically require the use of high amounts of chemicals and/or a significant energy input. A second important point of attention in algae cultivation is water recycle. Due to the low concentration of the algae in the cultivation (ca. 0,2 g/l dry algae in open ponds and 2 g/l in photobioreactors), large amounts of water need to be processed to produce algae paste. For example in open pond cultivation a production installation of 1000 ton dry algae per year requires about 700 m³/h water to be processed. Thus for large scale installation medium recycle is a sine qua non. A solution that tackles both issues simultaneously is the submerged flat panel membrane system. The membrane system is used as the first dewatering step in a hybrid system of algae harvesting with centrifugation as final concentration. This technology has the potential to lower energy and investment costs compared to centrifugation alone. The technology furthermore has major advantages on water recycling as > 95 % of the water needs to be removed to produce a 20 % paste of algae. As the membranes don't add any chemicals and remove all suspended solids and bacteria, the technology is very promising toward medium recycle. Submerged membrane filtration is preferred over other membrane filtration technologies (e.g. crossflow filtration), due to its low energy demand and low shearing forces. Membrane fouling is controlled by the cleaning effect of coarse bubbling aeration. Additionally, the flat panel membranes used in this study are backwashable (patented flat sheet membrane envelopes with an integrated permeate channel, IPC). The algae filtration experiments were performed on both lab and pilot-scale submerged reactors with different membranes (MF and UF), algae species (Nannochloropsis, Pavlova, Isochrysis, Phaeodactylum), algae concentration, filtration regimes and filtration cycle times and aeration flows. The results show that membrane operation with backwashing results in higher stable fluxes than operation with only relaxation which is the normal operating mode for submerged membranes. Moreover the flux of UF membranes is more stable than MF membranes and that shorter filtration times result in higher stable fluxes (in the backwashing regime). Aeration flow turned out to be critical towards achievable flux levels and the characteristics of the algae suspension. VITO will further develop and optimize the technology for harvesting and water recycle on pilot scale and demonstration scale. The latest information on the developments of this technology will be presented.

Immobilization of the enzymes plays a vital role in enhancing their applicability in a wide range of applications, thus ensuring the use of sustainable enzymatic processes over the conventional chemical processes on an industrial scale. This study provides the background information for the selection and screening of inorganic metal oxide ( MO) powders for their use as fillers in mixed matrix membranes for enzyme immobilization as the future aim. A total of 13 MOs, ranging in size from 0.01 mm to <5 mu m, were tested for their performance as a support for enzyme (beta-galactosidase) immobilization via adsorption. Alumina appeared to be the best performing MO with the amount and activity of the immobilized enzyme being 64 mg g(-1) and up to 288 U g(-1), respectively. The amount of immobilized enzyme on alumina (alpha-Al2O3 C and gamma-Al2O3) was > 3 times higher than ZrO2 (used as a reference MO in this study). Upon heat treatment at 900 degrees C, up to 15%, 52% and 42% decline was observed in the amount of immobilized enzyme in case of alumina metal oxides (MOs), ZrO2 and TiO2, respectively. The results suggested that both isoelectric point and surface area of the MO influence the immobilization. The most important observation in this study was that the bonding of the enzyme to the MO surface seems to be mediated by the bonding/interaction of the buffer to the enzyme.

We investigated if in-situ metal bioprecipitation (ISMP) is applicable to remediate a highly permeable zinc-contaminated aquifer at a metal-processing factory in Maasmechelen, Belgium. A large (more than 200m long and 70m wide) groundwater contamination plume has developed, with zinc concentrations in the range of 1-100mg/L, whereas the legal Flemish clean-up standard is 0.5mg/L. The estimated groundwater flow velocity is in the range 0.2-1m/d. The groundwater is relatively oxidized, naturally low in DOC (<1mg/L) and relatively low in sulfate (40-50mg/L). We conducted both laboratory feasibility tests as well as a long-term field pilot test in two sections of the plume. In the laboratory microcosm tests, zinc bioprecipitation (following addition of organic substrate and sulfate) removed more than 99% of the zinc from the water phase. Lactate, glycerol and vegetable oil were equally effective as substrates. 28-day anaerobic leaching tests indicated that the metal precipitates that were formed are stable, but they also suggested that substrate addition increases the solubility (leachability) of arsenic and manganese. In the field test, Zn concentrations were reduced by 2 to 3 orders of magnitude within the 232 day testing period and stayed low for the following 6 months in both pilot zones. In the field, no mobilization of arsenic occurred but manganese groundwater concentrations increased from 0.01-0.6mg/L to 0.4-6.5mg/L. Dissolved iron concentrations also increased markedly from below detection limits to concentrations as high as 67mg/L. Zinc concentrations in groundwater were closely correlated to pH and redox potential (Eh): plotting y=[Zn] against x=pH/log(Eh), an exponential relationship was found:

This paper consists of two parts – a detailed experimental investigation on the activity of supported threonine aldolase for use in a microreactor/flow reactor and a generic, explorative outline on potential productivity of enzymatic microreactors; with the need to use data from other enzymes here as well. Threonine aldolase was immobilized on different supports foreseen to be coated or packed within the micro- and millichannels of flow reactors. As enzyme supports, an innovative high-surface area and high-porosity silica (Nanosprings), the commercial Eupergit CM, and polymer membrane materials were used. For Eupergit CM, the share of chemically fixed enzymes and active enzymes were within the reported literature performance for enzyme immobilization. The values of immobilized enzymes on Nanosprings were somewhat lower. However, this is overcompensated by the larger specific surface area of Nanosprings and the availability of a new, second generation flow reactor concept, which involves a tight packing of stacked Nanosprings disks. For the investigated membranes, the density of immobilized enzymes was close to the reported value of capacity. From these experimental results, calculations on (maximally achievable) productivity of microreactors were made. This was combined with results of glucose oxidase immobilization on Nanosprings. The objective was to achieve high enzyme loadings to obtain reasonable space–time yields to ensure cost-effectiveness of the derived chemical reaction processes. These productivities were benchmarked to industrial production needs for two processes – a high-value pharmaceutical intermediate synthesis (towards chiral amino alcohols, using threonine aldolase) and a large-volumefine-chemical synthesis (the gluconic acid synthesis, using glucose oxidase), including suggestions for reactor scale-up. This shows that the present loading performance suffices in the case of high-value products, such as pharmaceuticals, but not for larger-volume, low-cost chemicals; at least under best-performance assumptions such as the avoidance of mass-transfer limitations.

β-Galactosidase is an important enzyme catalyzing not only the hydrolysis of lactose to the monosaccharides glucose and galactose but also the transgalactosylation reaction to produce galacto-oligosaccharides (GOS). In this study, β-galactosidase was immobilized by adsorption on a mixed-matrix membrane containing zirconium dioxide. The maximum β-galactosidase adsorbed on these membranes was 1.6 g/m2, however, maximal activity was achieved at an enzyme concentration of around 0.5 g/m2. The tests conducted to investigate the optimal immobilization parameters suggested that higher immobilization can be achieved under extreme parameters (pH and temperature) but the activity was not retained at such extreme operational parameters. The investigations on immobilized enzymes indicated that no real shift occurred in its optimal temperature after immobilization though the activity in case of immobilized enzyme was better retained at lower temperature (5 °C). A shift of 0.5 unit was observed in optimal pH after immobilization (pH 6.5 to 7). Perhaps the most striking results are the kinetic parameters of the immobilized enzyme; while the Michaelis constant (Km) value increased almost eight times compared to the free enzyme, the maximum enzyme velocity (Vmax) remained almost constant.

In situ bioprecipitation (ISBP), which involves immobilizing the metals as precipitates (mainly sulphides) in the solid phase, is an effective method of metal removal from contaminated groundwater. This study investigated the stability of metal precipitates formed after ISBP in two different solid-liquid matrices (artificial and natural). The artificial matrix consisted of sand, Zn (200 mg L(-1)), artificial groundwater and a carbon source (electron donor). Here the stability of the Zn precipitates was evaluated by manipulation of redox and pH. The natural system matrices included aquifer material and groundwater samples collected from three different metal (Zn and Co) contaminated sites and different carbon sources were provided as electron donors. In the natural matrices, metal precipitates stability was assessed by changing aquifer redox conditions, sequential extraction, and BIOMET assay. The results indicated that, in the artificial matrix, redox manipulation did not impact the Zn precipitates. However the sequential pH change proved detrimental, releasing 58% of the precipitated Zn back into liquid phase. In natural matrices, the applied carbon source largely affected the stability of metal precipitates. Elemental analysis performed on the precipitates formed in natural matrix showed that the main elements of the precipitates were sulphur with Zn and Co.

Organic carbon introduction in the soil to initiate remedial measures, nitrate infiltration due to agricultural practices or sulphate intrusion owing to industrial usage can influence the redox conditions and pH, thus affecting the mobility of heavy metals in soil and groundwater. This study reports the fate of Zn and Cd in sandy aquifers under a variety of plausible in-situ redox conditions that were induced by introduction of carbon and various electron acceptors in column experiments. Up to 100% Zn and Cd removal (from the liquid phase) was observed in all the four columns, however the mechanisms were different. Metal removal in column K1 (containing sulphate), was attributed to biological sulphate reduction and subsequent metal precipitation (as sulphides). In the presence of both nitrate and sulphate (K2), the former dominated the process, precipitating the heavy metals as hydroxides and/or carbonates. In the presence of sulphate, nitrate and supplemental iron (Fe(OH)3) (K3), metal removal was also due to precipitation as hydroxides and/or carbonates. In abiotic column, K4, (with supplemental iron (Fe(OH)3), but no nitrate), cation exchange with soil led to metal removal. The results obtained were modeled using the reactive transport model PHREEQC-2 to elucidate governing processes and to evaluate scenarios of organic carbon, sulphate and nitrate inputs.

About 45% of site contamination is caused by heavy metals. Metals are spread in the environment by mining activities, surface treatment and nonferrous processing. As heavy metals can not be degraded, the only existing riskreducing removal measures are removal or immobilization (leading to bioavailability reduction). Next to the often used but expensive pump and treat technologies, heavy metals can be immobilized by inducing sulfate reducing bacteria (SRB) to transform the sulfates that are very often present in groundwater (due to the metal mining or processing activities), into sulfides. These sulfides will precipitate the metals as insoluble metal sulfides. Several studies have demonstrated the feasibility of this In Situ Bioprecipitation Process (ISBP) for the removal of heavy metals from groundwater as well at lab scale (batch and column tests) as at field scale. However, some questions arise concerning the continuation of the process, and the efficiency and the sustainability of the precipitates. The study, based on more than 10 different studies, all done by the same authors, on different groundwaters and aquifer samples, will try to answer these questions. The presentation gives an overview of the guidelines necessary for a correct and successful bioprecipitation process with stable metal sulfide precipitates. It pays attention to the influence of the carbon source on the complexing of the metals and the efficiency of the induction of the bioprecipitation process, the Diels et al.: Heavy Metal Immobilization In Groundwater... Produced by The Berkeley Electronic Press, 2006 100 Contaminated SoilsHeavy Metals possible negative influence of acetate inhibition, the influence on the competence between sulfate reducers and methanogenic bacteria and the influence of low pH on the ISBP. The results will allow the correct implementation of the ISBP with an eye on the longevity and sustainability of the process and present the ISBP as a much more sustainable alternative to the pump and treat technology as a remediation measure for heavy-metals contaminated groundwaters.

Acetoacetyl-CoA reductase PhaB turned out to be capable of catalyzing the anti-Prelog asymmetric reduction of various β-ketoesters (1a-12a) and aromatic ketones (13a-29a). Particularly, PhaB showed high specific activity and excellent stereoselectivity (93.0%-99.8% eep) toward β-ketoesters. The specific activity of PhaB toward ethyl 4-chloroacetoacetate 11a was up to 1302.2 mU/mg. The molecular basis analysis showed the rotatable single bonds in β-ketoesters endow them with a high degree of structural flexibility and adaptability. Thus, β-ketoesters could quickly adjust their conformation and further form a productive conformation in the narrow substrate-binding pocket of the enzyme. In contrast, PhaB exhibited low specific activity and stereoselectivity toward the majority of aromatic ketones. The large steric hindrance and rigid structure resulted from aromatic rings made the aromatic ketones impossible to adjust their conformation as conveniently as β-ketoesters. Furthermore, it was found the halogen bond was the major driven force of the high specific activity of PhaB toward chlorinated β-ketoesters (10a and 11a), while the distribution of enzyme-substrate interactions was an important factor determining the enzyme activity besides the steric hindrance. Moreover, the geometric configuration of the substrate and the enzyme substrate-binding pocket played critical roles in determining the substrate binding mode and the enzyme stereoselectivity.