Continuous production of kyotorphin

The continuous alpha-chymotrypsin-catalyzed peptide synthesis of kyotorphin, tyrosyl-arginine, via the N(alpha) formyltyrosyl-arginine propyl ester is described. For continuous process development, two reaction systems were studied: immobilized alpha-chymotrypsin covalently bound to Eupergit C packed in a column, and soluble alpha-chymotrypsin utilizing an enzyme membrane reactor. Selectivities and kinetic parameters are discussed. The use of soluble enzyme in an enzyme membrane reactor proved superior to the covalently immobilized enzyme. A significant loss of enzyme activity and a certain decrease of selectivity was observed during immobilization. It was shown that the addition of organic solvent, in this case n-propanol, causes a severe diminuation of the enzyme activity.     

Nitrification-denitrification via nitrite accumulation for nitrogen removal from wastewaters

The biological nitrification-denitrification process is used extensively for removal of ammonia nitrogen from wastewaters. Saves in aeration, organic matter (for denitrification) and surplus sludge are achievable if nitrite accumulation is possible in the nitrification step. In this paper, operational parameters were studied for each process for maximum nitrite accumulation in the nitrification step and nitrite adaptation in the denitrification step. Nitrite accumulation during nitrification can be controlled by the dissolved oxygen (DO) concentration, presenting a maximum of 65% at around 0.7 mg DO/L. Denitrification can be adapted to nitrite and the process is stable if nitrite in the reactor is keep low. The performance of a continuous stirred tank reactor (CSTR) and an up flow sludge blanket reactor (USB) were compared. Once the operational parameters were established, a CSTR for nitrification and an USB reactor for denitrification were operated in series for 25 days. The process was stable and a steady state was maintained for 20 days, and 93.5% of overall nitrogen removal was achieved in the nitrification-denitrification via the nitrite process.     

Performance evaluation of reactors designed for bioconversion of wheat straw to animal feed

A chronology of reactor design from laboratory scale to pilot scale for the bioconversion of wheat straw to animal feed is presented. The engineering criteria considered at each stage of development are discussed. Designs were executed at each stage and their performance was compared based on engineering and bioconversion parameters. Illustrative detailed analyses of data obtained from performance evaluation experiments from selected designs are provided. Schematics diagrams of the different generations of reactor designs are also presented.     

Effect of modulation of enzyme inactivation on temperature optimization for reactor operation with chitin-immobilized lactase

Temperature is a critical variable to be optimized in any enzymatic process, producing opposite effects on enzyme activity and inactivation rate. Temperature functions for all kinetic and inactivation parameters were validated for chitin-immobilized yeast lactase (CIL). Enzyme inactivation was described by a two-stage series mechanism. The effect of galactose and lactose on inactivation was determined in terms of modulation factors that were positive for galactose and negative for lactose. Modulation factors were mild functions of temperature in the first stage and strong functions in the second stage of enzyme inactivation, where galactose positive modulation factors increase while lactose negative modulation factors decrease with temperature. Temperature-explicit functions for kinetic and inactivation parameters were incorporated into a scheme to optimize temperature in the simulation of a continuous packed-bed reactor operation with chitin-immobilized lactase, based on an annual cost objective function. Optimum temperature was 20°C at enzyme replacement of 25% residual activity, and increased only slightly at higher replacement frequencies. The effect of modulation factors on reactor design and temperature optimization is presented and discussed. Software for temperature optimization that allows the introduction of variations in all parameters and operational criteria to perform sensitivity analysis was developed.     

Performance of the continuous glucose isomerase reactor system for the production of fructose syrup

Glucose isomerase in the form of heat-treated whole-cell enzyme prepared from Streptomyces phaeochromogenus follows the reversible single-substrate reaction kinetics in isomerization of glucose to fructose. Based on the Kinetic constants determined and the mathematical model of the reactor system developed, the preformance of a plug-flow-type continuous-enzyme reactor system was studied experimentally and also simulated with the aid of a computer for the ultimate objective of optimization of the glucose isomerase reactor system. The enzyme decay function for both the enzyme storage and during the use in the continuous reactor, was found to follow the first-order decay kinetics. When the enzyme decay function is taken into consideration, the ideal homogeneous enzyme reactor kinetics provided a satisfactory working model without further complicatin of the mathematical model, and the results of computer simulation were found to be in good agreement with the experimental results. Under a given set of constraints the performance of the continuous glucose isomerase reactor system can be predicted by using the computer simulation method described in this paper. The important parameters studied for the optimization of reactor operation were enzyme loading, mean space time of the reactor, substrate feed concentration, enzyme decay constants, and the fractional conversion, in addition to the kinetic constants. All these parameters have significant effect on the productivity. Some unique properties of the glucose isomerization reaction and its effects on the performance of the continuous glucose isomerase reactor system have been studied and discussed. The reaction kinetics of glucose isomerase and the effects of both the enzyme loading and the changes in reaction rate within a continuous reactor on the productivity are all found to be of particular importance to this enzyme reactor system.     

Monitoring the impact of bioaugmentation on the start up of biological phosphorus removal in a laboratory scale activated sludge ecosystem

The acclimatisation of activated sludge to enhanced biological phosphorus removal (EBPR) conditions requires a period of about 40–100 days but its output remains hazardous. The impact of bioaugmentation on the start-up of a laboratory scale EBPR sequencing batch reactor was evaluated by process parameters measurement and microbial community dynamics monitoring using 16S rDNA targeted polymerase chain reaction-single strand conformation polymorphism electrophoresis (PCR-SSCP). Bioaugmentation: (1) speeded up the installation of good and stable EBPR in the bioaugmented reactor by about 15 days; (2) correlated with the transient enrichment of the sludge in the added microbial populations; and (3) favoured the long-term enrichment of the sludge in the phosphorus-accumulating organism (PAO) Candidatus Accumulibacter phosphatis. However, despite a lag time period, the control non-bioaugmented reactor ended up with comparable reactor parameters and microbial community evolution, suggesting that the same PAO populations were already present from the beginning in the original non-P-accumulating seed sludge. The potential of a true installation of the added microbial populations within the bioaugmented reactor compared to their substitution by indigenous similar populations is discussed. Competition between PAOs and the antagonistic glycogen accumulating organism Candidatus Competibacter phosphatis is also highlighted during EBPR start-up.     

Experimentelle Untersuchung,Modellierung und optimale Berechnung eines mehrstufigen Säulenfermentors II. Experimentelle Bestimmung der Parameter und Koeffizienten des mathematischen Modells

Using a system analysis for the investigation of all processes which occur in a biochemical reactor on the micro and macro level, a mathematical model was worked out. It characterizes the model of the kinetics and stoichiometry of the growth of microorganisms and the rules of hydrodynamics and mass transfer in form of blocks. Relating to the discussed mathematical total model [11], experimental data on which the calculation of the model parameters is based are described in this second part of the paper. They were determined not only directly from the cultivation process, but also from experiments with model media.     

Review of solid state fermentation for lignocellulolytic enzyme production: challenges for environmental applications

Within the context of increasing environmental concern, energy production from lignocellulosic substrates is gaining great interest. Enzymes have proven their efficiency in the degradation of the lignocellulosic complex but their use remains limited in environmental applications such as anaerobic digestion mainly due to their prohibitive cost. Therefore, solid state fermentation (SSF) emerges as an interesting alternative for the in situ production of lignocellulolytic enzymes. Various research efforts on the lab scale optimization of SSF are discussed. They are presented according to the type of inoculum used in the process: bacterial species and fungal species under both mesophilic and thermophilic conditions. In general, parameters that impact the SSF process include: substrate type and particle size, substrate pretreatment, inoculum, nutrient supplementation, moisture content, pH, aeration, temperature and mixing. Using different substrates, authors aim at maximizing enzyme production taking into account one to several of the indicated operational parameters. The reviewed research puts forward the adaptation of the operational parameters, enzyme production cost and loading, enzyme mixture quality and efficiency and finally reactor design as the main challenges for environmental large-scale application.     

Hydrolysis of oils by using immobilized lipase enzyme: A review

This review focuses on the use of immobilized lipase technology for the hydrolysis of oils. The importance of lipase catalyzed fat splitting process, the various immobilization procedures, kinetics, deactivation kinetics, New immobilized lipases for chiral resolution, reactor configurations, and process considerations are all reviewed and discussed.     

Startup of anaerobic fluidized bed reactors with acetic acid as the substrate

The startup of anaerobic fluidized bed reactors, which use Manville R-633 beads as the growth support media, acetate enriched bacterial culture as the inoculum, and acetic acid as the sole substrate, is studied. Tow startup strategies are evaluated: one based on maximum and stable substrate utilization and another based on maximum substrate loading controlled by reactor pH. The startup process is characterized using a number of operational parameters.The reactors again excellent total organic carbon (TOC) removal (i.e., > 97% at a feed concentration of 5000 mg TOC/L) and stable methane production (i.e., 0.90 L CH(4)/g TOC, where TOC(r) is TOC removed) at a early stage of the startup process, regardless of the strategies applied. The loading can be increased rapidly without the danger of being overloaded. Significant losses of growth support media and biomass caused by gas effervescence at higher loadings limits the maximum loading that can be safely applied during startup process.A high reactor immobilized biomass inventory is achievable using the porous growth support media (e.g., Manville 633 beads). A rapid increase in loading creates a substrate rich environment that yields more viable reactor biomass. Both substrate utilization rate (batch and continuous) and immobilized biomass inventory stabilize concomitantly at the late stage of the startup process, indicating the attainment of steady-state conditions in reactors. Therefore, they are better parameters that TOC removal and methane production for characterizing the entire startup process of aerobic fluidized bed reactor.The strategy based on maximum substrate loading controlled by reactor pH significantly shortens the startup time. In this case, the reactor attains steady-state conditions approximately 140 days after startup. On the other hand, a startup time of 200 days is required when the strategy based maximum substrate utilization is adopted. (c) 1993 John Wiley & Sons, Inc.     

Synthesis of protective antigens during submerged cultivation of Vibrio cholerae

The effectiveness of dot immunoanalysis for evaluating the dynamics of the synthesis of O-antigen, cholera toxin, neuraminidase, adhesin CFA1 in the process of the reactor cultivation of V. cholerae used for the production of oral chemical cholera vaccine is shown. The established regularities of the synthesis of the protective antigens of V. cholerae in the process of scaled-up cultivation are discussed.     

Mathematical and kinetic modeling of biofilm reactor based on ant colony optimization

Inverse estimation of model parameters via mathematical modeling route, known as inverse modeling (IM), is an attractive alternative approach to the experimental methods. This approach makes use of efficient optimization techniques in the course of solution of an inverse problem with the aid of measured data. In this study, a novel optimization method based on ant colony optimization (ACO), denoted by ACO-IM, is presented for inverse estimation of kinetic and film thickness parameters of biofilm models that describe an experimental fixed bed anaerobic reactor. The proposed optimization method for parameter estimation emulates the fact that ants are capable of finding the shortest path from a food source to their nest by depositing a trial of pheromone during their walk. The efficacy of the ACO-IM for numerical estimation of bio-kinetic parameters is demonstrated through its application for the anaerobic treatment of industry wastewater in a fixed bed biofilm process. The results explain the rigorousness of mathematical models, the form of kinetic and film thickness models and the type of packing to be used with the biofilm process for accurate determination of kinetic and film thickness parameters so as to ensure reliable predictive performance of the biofilm reactor models.     

Ecological study of a bioaugmentation failure

A nitrifying sequencing batch reactor was inoculated twice with the aerobic denitrifying bacterium Microvirgula aerodenitrificans and fed with acetate. No improvement was obtained on nitrogen removal. The second more massive inoculation was even followed by a nitrification breakdown, while at the same time, nitrification remained stable in a second reactor operated under the same conditions without bioaugmentation. Fluorescent in situ hybridization with rRNA-targeted probes revealed that the added bacteria almost disappeared from the reactor within 2 days, and that digestive vacuoles of protozoa gave strong hybridization signals with the M. aerodenitrificans -specific probe. An overgrowth of protozoa, coincident with the disappearance of free-living bacteria, was monitored by radioactive dot-blot hybridization only in the bioaugmented reactor. Population dynamics were analysed with a newly developed in situ quantification procedure of the probe-targeted bacteria. The nitrifying groups of bacteria decreased in a similar way in the bioaugmented and non-bioaugmented reactors. Other bacterial groups evolved differently. The involvement of different ecological parameters are discussed separately for each reactor. These results underline the importance of predator–prey interaction and illustrate the undesirable effects of massive bioaugmentation.     

Nanobiotechnology as a novel paradigm for enzyme immobilisation and stabilisation with potential applications in biodiesel production

Nanobiotechnology is emerging as a new frontier of biotechnology. The potential applications of nanobiotechnology in bioenergy and biosensors have encouraged researchers in recent years to investigate new novel nanoscaffolds to build robust nanobiocatalytic systems. Enzymes, mainly hydrolytic class of enzyme, have been extensively immobilised on nanoscaffold support for long-term stabilisation by enhancing thermal, operational and storage catalytic potential. In the present report, novel nanoscaffold variants employed in the recent past for enzyme immobilisation, namely nanoparticles, nanofibres, nanotubes, nanopores, nanosheets and nanocomposites, are discussed in the context of lipase-mediated nanobiocatalysis. These nanocarriers have an inherently large surface area that leads to high enzyme loading and consequently high volumetric enzyme activity. Due to their high tensile strengths, nanoscale materials are often robust and resistant to breakage through mechanical shear in the running reactor making them suitable for multiple reuses. The optimisation of various nanosupports process parameters, such as the enzyme type and selection of suitable immobilisation method may help lead to the development of an efficient enzyme reactor. This might in turn offer a potential platform for exploring other enzymes for the development of stable nanobiocatalytic systems, which could help to address global environmental issues by facilitating the production of green energy. The successful validation of the feasibility of nanobiocatalysis for biodiesel production represents the beginning of a new field of research. The economic hurdles inherent in viably scaling nanobiocatalysts from a lab-scale to industrial biodiesel production are also discussed.     

Two-phase anaerobic digestion for production of hydrogen-methane mixtures

An anaerobic digestion process to produce hydrogen and methane in two sequential stages was investigated, using two bioreactors of 2 and 15 L working volume, respectively. This relative volume ratio (and shorter retention time in the second, CH(4)-producing reactor) was selected, in part, to test the assumption that separation of phase can enhance metabolism in the second methane producing reactor. The reactor system was seeded with conventional anaerobic digester sludge, fed with a glucose-yeast extract--peptone medium and operated under conditions of relatively low mixing, to simulate full scale operation. A total of nine steady states were investigated, spanning a range of feed concentrations, dilution rates, feed carbon to nitrogen ratios and degree of integration of the two stages. The performance of this two-stage process and potential practical applications for the production of clean-burning hydrogen-methane mixtures are discussed.     

Generalized rate equation for one-substrate enzymatic reactions

A generalized rate equation for formally two-step enzymatic reaction has been derived. In this derivation the reaction rate has been regarded as a function of substrate conversion. Applications of this equation to reactor operating parameters and productivity calculations are presented and discussed.     

Comparison of the Baker's yeast process performance in laboratory and production scale

A 215 m³ industrial bubble column reactor for fedbatch production of Baker's yeast was sampled for sugar, to investigate the extent of concentration gradients. The results verify that such gradients exist: the concentration is higher closer to the feeding point. Effects of sugar heterogeneities at different scales were studied by 1)?performing a volumetric scale-down of the industrial process in a laboratory stirred tank reactor (STR); 2) performing the same scaled down process in a Scale-Down Reactor (SDR) with repeated short term exposure of the cells to high sugar concentrations. In this reactor about 10% of the Baker's yeast culture was intermittently exposed to high (0.45–1.9?g?l^?1) concentrations of sugar, for periods of 60 seconds. It was found that physiological parameters of glycolysis and respiration were affected by environmental heterogeneities: 1) A biomass yield reduction of about 6–7% was found, with both the production reactor and the SDR, as compared to the homogeneous reactor. The loss of yield is interpreted in terms of a metabolic by-pass via ethanol, where cells are consuming and producing ethanol with different yields. 2) The maximum respiration rate was higher in cells produced in the production unit and in the SDR. 3) The product quality, expressed as gassing power of the yeast in a dough, was increased for sweet and non-sugar doughs in the SDR, and for sweet doughs in the production reactor. Thus, the SDR, when run with defined glucose gradients, in some aspects resembles the large reactor. It could be regarded as a tool for scale-down and scale-up studies and may be useful in investigations on the scale-up sensitivity of a process.     

Biodegradation of di-n-butyl phthalate (DnBP) in bioaugmented bioslurry phase reactor

Bioremediation of di-n-butyl phthalate (DnBP) in soil was studied with various concentrations in a bioslurry phase batch reactor operated in sequenting batch mode (bioaugmented with effluent treatment plant (ETP) microflora) for a total cycle period of 96h. Process performance during the reactor operation was assessed by monitoring DnBP concentration and biochemical process parameters viz., pH, dissolved oxygen (DO), colony forming units (CFU) and oxygen uptake rate (OUR), during the sequence phase operation. The degradation rate was observed to be rapid at lower substrate concentrations and found to be slow as the substrate concentration increased. The potent bacterial strain was also isolated from the slurry phase reactor. Metabolites formed during the degradation of DnBP in the slurry phase reactor were identified. Studies on the kinetics and half-life of the reaction revealed that the degradation process followed zero-order kinetic model.     

Cost optimization of activated sludge systems

Activated sludge is a widely used aerobic biological waste-water treatment process. A rational approach to least cost design of an integrated system is described which includes the following processes: activated sludge reactor, final settling tanks, gravity thickening, and aerobic sludge digestion. Both capital and operation and maintenance costs are considered. Biological reactor design is based on microbial kinetic concepts and continuous culture of microorganisms theory. Biological solids retention time (θ_c) is utilized as the primary independent design variable to which system performance is related, e.g., effluent quality, ammonia oxidation, and excess sludge production. Liquid-biomass separation is based on the batch flux technique, a rational approach to design of gravity separators (final settling tanks). Trade-offs among reactor volume, clarifier size, recycle pumping capacity, thickener capacity, digester volume, air requirements, and sludge production are discussed. The optimum design is taken as the combination of these parameters within the acceptable design domain, determined by effluent quality criteria, that results in minimum cost. While the method described is general, design of a given treatment system depends on availability, from lab or pilot studies, of system specific numerical values for biological growth coefficients and biomass setting characteristics. A design example illustrates the approach.     

Lactic acid production by immobilized Lactobacillus casei in recycle batch reactor: a step towards optimization.

Different nutritional and process parameters influencing lactic acid production by Lactobacillus casei, adsorbed to Poraver beads in a recycle batch reactor system, were studied in an attempt to set up a system having a long operational lifetime and permitting use of high substrate concentrations for maximal conversion to the product. The presence of lactose, even as a minor fraction of the total sugar amount, was necessary for complete utilization by the organism for growth and conversion to lactate. Hydrolysed whey protein constituted a richer source of nitrogen compared to yeast extract. Addition of lactate to the medium at the start of the process resulted in severe inhibition compared with the normal process. For a homofermentative process, pH 6.0 was found to be optimal. The overall productivity of the recycle system was higher under all conditions studied in comparison with the batch process using free cells. Enhancement in productivity in the recycle batch reactor was also accompanied by an increase in density of suspended cells. However, the contribution of the suspended cells to the overall reactor productivity was not noticeable. The bead size of the matrix was found to be important for operational stability of the reactor.