Characterization of a pilot plant airlift tower loop bioreactor. I: evaluation of the phase properties with model media

Investigations were carried out in a 9 m high, 4 m(3) volume, pilot plant airlift tower loop bioreactor with a draft tube. The reactor was characterized by measuring residence time distributions of the gas phase using pseudostochastic tracer signals and a mass spectrometer and by evaluating the mixing in the liquid phase with single-pulse tracer inputs. The local gas holdup and the bubble size (piercing length) were measured with two-channel electrical conductivity probes. The mean residence times and the intensities of the axial mixing in the riser and downcomer and the circulation times of the phases as well as the fraction of the recirculated gas phase were evaluated. The gas holdup in the riser is nearly uniform along the reactor. In the downcomer, it diminishes from top to bottom. The liquid phase dispersion coefficients, D(L), are smaller than those measured in the corresponding bubble columns. In the pilot plant with tap water the following relationship was found: D(Lr) = cw(SG) (n); with c = 203.4; n = 0.5;D(Lr)(cm(2) s(-1);) and W(SG)(cm s(-1)) where D(Lr) is the longitudinal dispersion coefficient in the riser and W(SG) is the superficial gas velocity. The gas phase dispersion coefficients in the riser of the pilot plant, D(Gr), are also enlarged with increasing superficial gas velocity, W(SG), however, no simple relationship exists. Parameter D(Gr) is the highest in the presence of antifoam agents, intermediate in tap water, and the smallest in ethanol solution.     

Characterization of a pilot plant airlift tower loop reactor: III. Evaluation of local properties of the dispersed gas phase during yeast cultivation and in model media

The local properties of the dispersed gas phase (gasholdup, bubble diamater, and bubble velocity) were measured and evaluated at different positions in the riser and downcomer of a pilot plant reactor and, for comparison, in a laboratory reactor. These were described in Parts I and II of this series of articles during yeast cultivation and with model media. In the riser of the pilot plant reactor, the local gas holdup and bubble velocities varied only slightly in axial direction. The gas holdup increased considerably, while the bubble velocity increased only slightly with aeration rate. The bubble size diminished with increasing distance from the aerator in the riser, since the primary bubble size was larger than the equilibrium bubble size. In the downcomer, the mean bubble size was smaller than in the riser. The mean bubble size varied only slightly, the bubble velocity was accelerated, and the gas holdup decreased from top to bottom in the downcomer. In pilot plant at constant aeration rate, the properties of the dispersed phase were nearly constant during the batch cultivation, i.e., they depended only slightly on the cell concentration. In the laboratory reactor, the mean bubble sizes were much larger than in the pilot plant reactor. In the laboratory reactor, the bubble velocities in the riser and downcomer increased, and the mean gas holdup and bubble diameter in the downcomer remained constant as the aeration rate was increased.     

Novel method for cell immobilization and its application for production of oligosaccharides from sucrose

AIMS: The purpose of the present investigation was to develop a novel method for cell immobilization. METHODS AND RESULTS: Aureobasidium pullulans cells were mixed with an alginate solution, and the mixture was extruded to form small gel beads as hydrated-immobilized cells. The beads were then placed at -15 degrees C for 6-24 h to induce freeze-dehydration. The freeze-dehydration resulted in shrinkage of beads as a result of water removal reducing bead volume by 82% and bead weight by 85%. The dehydrated beads were successfully used for the production of fructo-oligosaccharides in a model reactor system. CONCLUSIONS: Dehydrated beads may provide some commercial advantages over conventional immobilized cells. SIGNIFICANCE AND IMPACT OF THE STUDY: This study shows that bioreactor performance can be improved up to two times by the use of the dehydrated beads.     

The effect of bioreactor configuration on production of HIV and cell-virus interaction

In an attempt to establish a bioreactor system for generation of HIV that is practicable, efficient, biologically contained, and capable of scale up, the production of two strains of this virus was examined in suspension culture and the Porosphere fixed bed system. HIV 1 and HIV 2 were grown successfully in both these types of reactor. The porosphere reactor theoretically appears to offer a better environment for HIV production, but evidence for significantly improved yields from this system, compared to suspension, was equivocal. However, this configuration facilitated media changes during culture. The data clearly showed that the culture system and cell environment significantly affected cell-virus interrelationships. Switches between lytic — and persistent — type infections, and changes in the virus population were observed.     

Oxygen transfer in a pulse bioreactor

Oxygen transfer in a novel pulse bioreactor has been evaluated. The agitator consists of a series of alternately fixed and movable parallel plates mounted so that the movable plates vibrate at 30 Hz causing a pulsating fluid motion. Pure oxygen, at pressures up to 5 atm, diffuses through silicone rubber tubing that also vibrates at 30 or 60 Hz. The main feature of this bioreactor is high oxygen transfer with low shear to prevent damage to fragile animal cell membranes. We estimate that sufficient oxygen can be supplied to support over 10(8) cells/mL of human diploid foreskin cells growing on microcarriers. (c) 1993 John Wiley & Sons, Inc.     

Monoclonal antibody production: viability improvement of RC1 hybridoma cell in different types of bioreactor

The study was done to improve the viability of the RC1 hybridoma cell in order to produce more amount of monoclonal antibody (mAb). By using the optimized media, the cell had been cultured in two bioreactor systems which were the MiniPerm and Stirred Tank bioreactor (ST bioreactor), and the results were compared to the one obtained by using the T-Flask bioreactor which was used as a standard. The results showed that the ST bioreactor was able to improve the viability of the cell to the value of 91.8% which was a little bit better than the one obtained by the MiniPerm bioreactor (88.6%) and far better than that of achieved by the T-Flask bioreactor (76.4%). This was well correlated with the good growth performance of the cell in the ST bioreactor with the specific growth rate (μ) value of 0.0289 h⁻¹ followed by MiniPerm bioreactor with the value of 0.0243 h⁻¹ and then the T-Flask with the value of 0.0151 h⁻¹. The low value of doubling time (t _d ) obtained in the ST bioreactor (24 h) compared to the one obtained in the MiniPerm (29 h) and T-Flask bioreactor (46 h) had also contributed to the higher value of cell viability. As a result a higher concentration of mAb was able to be produced by the ST bioreactor (0.42 g l⁻¹) compared to that of the MiniPerm (0.37 g l⁻¹) and T-Flask bioreactor (0.23 g l⁻¹).     

Production of Ethyl (R)-2-Hydroxy-4-phenylbutanoate via Reduction of Ethyl 2-Oxo-4-phenylbutanoate in an Interface Bioreactor

Ethyl (R)-2-hydroxy-4-phenylbutanoate [(R)-EHPB], a useful intermediate for the synthesis of various anti-hypertension drugs, was produced via microbial reduction of ethyl 2-oxo-4-phenylbutanoate [EOPB] in an interface bioreactor. Rhodotorula minuta IFO 0920 and Candida holmii KPY 12402 were selected as the best type culture and isolated yeasts, respectively. The highest enantiomeric excess of (R)-EHPB produced by R. minuta and C. holmii were 95 and 94%, respectively. C. holmii was used for the reduction of EOPB in a pad-packed interface bioreactor (inner volume, 3 liter). After incubation for 4 days, 4.4 g of (R)-EHPB was obtained via extraction with methanol followed by column chromatography. The overall yield, chemical purity, and enantiomeric excess of (R)-EHPB were 58%, 99.1%, and 90%, respectively.     

Liquid-phase mass transfer coefficients in bioreactors

Liquid-phase mass transfer coefficient in bioreactors have been examined. A theoretical model based on the surface renewal concept has been devloped. The predicted liquid-phase mass transfer coefficients are compared with the experimental data for a mycelial fermentation broth (Chaetomium cellulolyticum) and model media (carboxymethyl cellulose) in a bench-scale bubble column reactor. The liquid-phase mass transfer coefficient is evaluated by dividing the volumetric mass transfer coefficient obtained experimentally by the specific surface area estimated using the available correlations. The available literature data in bubble column and stirred tank bioreactors is also used to test the validity of the proposed model. A reasonable agreement between the model and the experimental data is found.     

A fiber-bed bioreactor for anchorage-dependent animal cell cultures: part II. Scaleup potential

A simple hydrodynamic model is introduced to describe the airlift fiber-bed bioreactor, which can enhance the volumetric productivity of anchorage-dependent animal cell cultures. By applying the model, liquid flow rates and volumetric mass transfer coefficients are predicted and are in agreement with experimental measurements. Consequently, the optimal reactor configuration giving the maximal oxygen supply is derived. Also, theoretical scaleup potential of this concentric internal loop reactor is considered for volumes ranging from 10 to 67,000 L with which cell densities of 5.1 x 10(7) and 1.2 x 10(7) cells/cm(3), respectively, can be maintained.     

植物生物反应器研究现状、瓶颈及策略

近10年,植物作为重组蛋白生产系统是生命科学中研究最活跃领域之一。植物系统具有低成本、安全和易规模化优势,其表达生物活性药用蛋白能力已被许多研究所证实;同时,植物药用蛋白产品还表现出潜在的市场和广阔应用前景。鉴于此,回顾了植物生物反应器兴起,介绍了植物表达系统和重组蛋白研究现状,综述了植物生物反应器面临瓶颈问题、解决对策和未来一段时间内研究热点;在展望植物生物反应器前景同时,对我国研究现状、与国外差距和未来发展应采取策略进行了讨论。     

Conversion of Orotic Acid to Uridine-5′-monophosphate by Enzymes of Micrococcus glutamicus

An enzyme system which could convert orotic acid to uridine-5′-monophosphate (5′-UMP) was found in cell-free extract of a threonine-requiring auxotroph of Micrococcus glutamicus (Syn. Corynebacterium glutamicum) 534 Co-147. This reaction required 5-phosphoribosylpyrophosphate (PRPP) and magnesium ion as essential components. The product of the enzyme reaction was separated by ion exchange resin chromatography and identified to be uridine-5′-monopbosphate. From the stoichiometric studies and other characteristics, it became evident that this enzyme reaction proceeded according to the following equation and was assumed to be catalyzed by orotidine-5′-monophosphate pyrophosphorylase and orotidine-5′-monophosphate decarboxylase. $\text{Orotic}\text{ acid + PRPP}\underset{{\text{Mg}}^{++}}{\to }{5}^{\prime }?\text{UMP}+\text{PPi}+{\text{CO}}_2$     

Regeneration of plants using nutrient mist culture

Summary A nutrient mist was used forin vitro culture of plant tissue in a novel bioreactor, wherein the tissues were grown on a biologically inert screen within a sterile chamber which allows excess media to drain away from the tissue. Plants tested includedDaucus, Lycopersicon, Ficus, Cinchona, andBrassica. The latter 4 genera were fully regenerated within the bioreactor. Tissue inocula included callus, anthers, and shoot meristems. All plants grew at least as well in nutrient mists as in agar and always produced a greater quantity of shoots of a higher quality and often faster than agar cultures. Cost analysis estimates showed up to a 65% savings in production costs (labor and materials) could be realized using nutrient mist culture. Nutrient mist culture offers significant improvements in the micropropagation of plants.     

Synchronized mammalian cell culture: Part I—A physical strategy for synchronized cultivation under physiological conditions

Conventional analysis and optimization procedures of mammalian cell culture processes mostly treat the culture as a homogeneous population. Hence, the focus is on cell physiology and metabolism, cell line development, and process control strategy. Impact on cultivations caused by potential variations in cellular properties between different subpopulations, however, has not yet been evaluated systematically. One main cause for the formation of such subpopulations is the progress of all cells through the cell cycle. The interaction of potential cell cycle specific variations in the cell behavior with large‐scale process conditions can be optimally determined by means of (partially) synchronized cultivations, with subsequent population resolved model analysis. Therefore, it is desirable to synchronize a culture with minimal perturbation, which is possible with different yield and quality using physical selection methods, but not with frequently used chemical or whole‐culture methods. Conventional nonsynchronizing methods with subsequent cell‐specific, for example, flow cytometric analysis, can only resolve cell‐limited effects of the cell cycle. In this work, we demonstrate countercurrent‐flow centrifugal elutriation as a useful physical method to enrich mammalian cell populations within different phases of a cell cycle, which can be further cultivated for synchronized growth in bioreactors under physiological conditions. The presented combined approach contrasts with other physical selection methods especially with respect to the achievable yield, which makes it suitable for bioreactor scale cultivations. As shown with two industrial cell lines (CHO‐K1 and human AGE1.HN), synchronous inocula can be obtained with overall synchrony degrees of up to 82% in the G1 phase, 53% in the S phase and 60% in the phase, with enrichment factors ( ) of 1.71, 1.79, and 4.24 respectively. Cells are able to grow with synchrony in bioreactors over several cell cycles. This strategy, combined with population‐resolved model analysis and parameter extraction as described in the accompanying paper, offers new possibilities for studies of cell lines and processes at levels of cell cycle and population under physiological conditions. © 2014 American Institute of Chemical Engineers Biotechnol. Prog., 31:165–174, 2015     

A fiber-bed bioreactor for anchorage-dependent animal cell cultures: part I. Bioreactor design and operations

A concentric-cylinder airlift reactor, in which the annulus is a packed bed of glass fibers, has been developed in order to facilitate the scaleup and enhance the volumetric productivity of anchorage-dependent animal cell cultures. In this bio-reactor, oxygen-containing gas is sparged through the inner draft tube, causing bubble-free medium to flow through the fiber bed in the outer cylinder and providing both oxygenation and convective nutrient transfer to the cells. Several other desirable features for reactor operation are also provided by this design. Cell cultivations in this bioreactor have been successfully carried out and provide data for the feasibility of the large-scale cell cultivation.     

Comparison of cell growth in T-flasks, in micro hollow fiber bioreactors, and in an industrial scale hollow fiber bioreactor system

In this article, cell growth in a novel micro hollow fiberbioreactor was compared to that in a T-flask and theAcuSyst-Maximizer®, a large scale industrial hollowfiber bioreactor system. In T-flasks, there was relativelylittle difference in the growth rates of one murine hybridomacultured in three different media and for three other murinehybridomas cultured in one medium. However, substantialdifferences were seen in the growth rates of cells in themicro bioreactor under these same conditions. These differencecorrelated well with the corresponding rates of initial cellexpansion in the Maximizer. Quantitative prediction of thesteady-state antibody production rate in the Maximizer was moreproblematic. However, conditions which lead to faster initialcell growth and higher viable cell densities in the microbioreactor correlated with better performance of a cell line inthe Maximizer. These results demonstrate that the microbioreactor is more useful than a T-flask for determining optimalconditions for cell growth in a large scale hollow fiberbioreactor system.     

Industrial wastewater bioreactors: sources of novel microorganisms for biotechnology

Microorganisms exist in nature as members of complex, mixed communities. The microbial communities in industrial wastewater bioreactors can be used as model systems to study the evolution of new metabolic pathways in natural ecosystems. The evolution of microbial metabolic capability in these bioreactors is presumably analogous to phenomena that occur in natural ecosystems. The microorganisms in these bioreactors compete for different carbon sources and constantly have to evolve new metabolic capabilities for survival. Thus, industrial bioreactors should be a rich source of novel biocatalysts.     

Development of a predictive description of an immobilized-cell, three-phase, fluidized-bed bioreactor

A Large bioreactor is an inhomogenous system with concentration gradients which depend on the fluid dynamics and the mass transfer of the reactor, the feeding strategy, the saturation constant, and the cell density. The responses of Escherichia coli cells to short-term oscillations of the carbon/energy substrate in glucose limited fed-batch cultivations were studied in a two-compartment reactor system consisting of a stirred tank reactor (STR) and an aerated plug flow reactor (PFR) as a recycle loop. Short-term glucose excess or starvation in the PFR was simulated by feeding of glucose to the PFR or to the STR alternatively. The cellular response to repeated short-term glucose excess was a transient increase of glucose consumption and acetate formation. But, there was no accumulation of acetate in the culture, because it was consumed in the STR part where the glucose concentration was growth limiting. However, acetate accumulated during the cultivation if the oxygen supply in the PFR was insufficient, causing higher acetate formation. The biomass yield was then negatively influenced, which was also the case if the PFR was used to simulate a glucose starvation zone. The results suggest that short-term heterogeneities influence the cellular physiology and growth, and can be of major importance for the process performance. (c) 1995 John Wiley & Sons, Inc.     

Biodegradation of Phenol Using Immobilized Nocardia hydrocarbonoxydans in a Pulsed Plate Bioreactor: Effect of Packed Stages,Cell Carrier Loading,and Cell Acclimatization on Startup and Steady-State Behavior

The effect of the number of stages and cell carrier loading on the steady-state and startup performance of a continuous pulsed plate bioreactor with glass beads as the cell carrier material for biodegradation of phenol in wastewater using immobilized Nocardia hydrocarbonoxydans has been studied. It was found that the performance of the pulsed plate bioreactor during startup and at steady state can be improved by an increase in cell carrier loading, number of stages, total plate stack height, and with a decrease in plate spacing. The startup time for the continuous bioreactor can be decreased by increasing the number of preacclimatization steps for the cells. The attainment of steady effluent phenol concentration can be considered as an indication of steady state of the continuous bioreactor, as when phenol concentration attained a steady value, biofilm thickness, and the attached biomass dry weight also attained a constant value.     

The effects on operation conditions of sludge retention time and carbon/nitrogen ratio in an intermittently aerated membrane bioreactor (IAMBR)

An intermittently aerated membrane bioreactor (IAMBR) system has been developed to improve the efficiency of nutrient removal, and for the stable treatment of organic matter which is contained as suspended solid (SS) in the influent. The important operating factors of an intermittently aerated bioreactor (IABR) are sludge retention times (SRTs) and carbon/nitrogen (C/N) ratios. Because research on IAMBR is young, this paper explores the effect of SRTs and C/N ratios on these systems. For SRTs of 20, 25, 30, and 40 days, there was little difference in the removal of COD, T–N, and T–P. In comparing C/N ratios of 4.5, 7, and 10, the COD concentration in permeate with a C/N ratio of 10 was most stable, although the concentration of organic matter in the influent was high. The removal efficiencies of T–N and T–P in permeate with a C/N ratio of 10 were the highest at 92.9% and 88.9%, respectively. This implies that a C/N ratio above 10 should be maintained for a nutrient removal efficiency of approximately 90%.     

A novel biphasic extractive membrane bioreactor for minimization of membrane-attached biofilms

Extractive membrane bioreactor (EMB) systems offer a means of biologically treating wastewaters, but, like other membrane processes, are constrained by their tendency to be fouled by membrane-attached biofilms (MABs). This study describes a new approach to eradicate MAB formation and accumulation in EMB systems. To this end, an innovative EMB configuration, the biphasic extractive membrane bioreactor (BEMB), has been developed. In BEMB systems, the two main constituents of the EMB process, membrane and bacteria, are kept separated and interact via a suitable recirculating solvent. Nineteen candidate solvents were tested to assess their suitability for BEMB application. Based on the results of the solvent selection, guidelines are provided to screen solvents for BEMB application. BEMB and EMB runs were carried out to demonstrate the effectiveness of BEMB technology in avoiding MAB accumulation and to compare BEMB and EMB performance. A synthetic wastewater containing monochlorobenzene (MCB) was used as a model system. Abiotic BEMB and EMB runs were carried out and used as comparative references for estimating the effect of MAB accumulation on system performance. MAB thickness in the BEMB systems was controlled at 18 microm during 1 month of operation, whereas, in the EMB systems, MAB thickness reached 1250 microm. Analysis of mass transport in EMB and BEMB systems revealed that the high affinity of the permeating molecules for the solvent may contribute to a reduction in shell-side mass transfer resistance. This reduction of shell-side mass transfer resistance and the absence of MAB accumulation led to overall mass transfer coefficients of about sevenfold greater (4.5 x 10(-5) m s(-1)) in the BEMB system than in the EMB system (0.6 x 10(-5) m s(-1)).