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  • Articles  (113)
  • immobilization  (50)
  • bioreactor  (35)
  • continuous culture  (29)
  • 1995-1999  (113)
  • 1980-1984
  • 1925-1929
  • Process Engineering, Biotechnology, Nutrition Technology  (113)
  • 1
    Electronic Resource
    Electronic Resource
    Kinetics of lactose transport inKluyveromyces fragilis grown in a chemostat on diluted whey permeate (1995)
    Springer
    Journal of industrial microbiology and biotechnology 15 (1995), S. 45-48 
    Details
    ISSN: 1476-5535
    Keywords: Kluyveromyces fragilis ; lactose transport ; continuous culture
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Lactose transport was studied inKluyveromyces fragilis grown in lactose-limited chemostat cultures. Kinetic parameters were determined using a method based on genetic population evolution. Lactose transport was carried out via three carriers characterized respectively byK m of 0.1 mM, 3 mM and 15.5 mM. The synthesis of these lactose carriers and their capacity (V max) are dependent on the dilution rate (D). At D=0.12 h−1, the high affinity transporter is prominent. For intermediate dilution rate, only the high and the medium affinity systems are present. In cells growing at D=0.4 h−1, these carriers are absent but instead, the low affinity transporter is present. The effect on lactose transport of such metabolic inhibitors as CCCP, a proton ionophore, and Antimycin A, an energy inhibitor, were also investigated. The high affinity system is the most sensitive to the effect of these inhibitors. Lactose transport through this carrier is probably a mechanism dependent on the proton motive force.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF01570012
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  • 2
    Electronic Resource
    Electronic Resource
    Effect of oxygen on denitrification in continuous chemostat culture withComamonas sp SGLY2 (1996)
    Springer
    Journal of industrial microbiology and biotechnology 16 (1996), S. 124-128 
    Details
    ISSN: 1476-5535
    Keywords: denitrification ; continuous culture ; oxygen ; Comamonas sp
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Continuous cultures ofComamonas sp SGLY2 were grown anaerobically prior to establishing steady states at different oxygen flow rates. At a low oxygen transfer rate, no dissolved oxygen accumulated in the medium and all nitrate was reduced to dinitrogen. Concurrently with the increase of dissolved oxygen concentration in the liquid phase, the rate of denitrification decreased. However, at a dissolved oxygen concentration near saturation (33 mg L−1), a part of the electron flow always diverted to nitrate with production of dinitrogen: the aerobic denitrification rate was equivalent to 35% of that calculated under anaerobic conditions. These experiments reflected the co-utilization of oxygen and N-oxides and the production of dinitrogen, up to saturated conditions, which implied synthesis and activity of the four denitrifying enzymes under various aeration conditions.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF01570072
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  • 3
    Electronic Resource
    Electronic Resource
    Environmental applications of immobilized microbial cells: A review (1996)
    Springer
    Journal of industrial microbiology and biotechnology 16 (1996), S. 79-101 
    Details
    ISSN: 1476-5535
    Keywords: alginate ; bacteria ; biodegradation ; bioremediation ; κ-carrageenan ; encapsulation ; immobilization ; microorganisms ; soil
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Immobilized microbial cells have been used extensively in various industrial and scientific endeavours. However, immobilized cells have not been used widely for environmental applications. This review examines many of the scientific and technical aspects involved in using immobilized microbial cells in environmental applications, with a particular focus on cells encapsulated in biopolymer gels. Some advantages and limitations of using immobilized cells in bioreactor studies are also discussed.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF01570068
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  • 4
    Electronic Resource
    Electronic Resource
    Comparison of citric acid production byAspergillus niger immobilized in gels and cryogels of polyacrylamide (1996)
    Springer
    Journal of industrial microbiology and biotechnology 16 (1996), S. 351-353 
    Details
    ISSN: 1476-5535
    Keywords: citric acid ; Aspergillus niger ; immobilization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Aspergillus niger was immobilized in cryogels and in conventional gels of polyacrylamide. The growth of cells entrapped in two kinds of gels and the production of citric acid by the immobilized cells were investigated and compared. Cells immobilized in cryogels were more suitable for citric acid production.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF01570114
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  • 5
    Electronic Resource
    Electronic Resource
    Analysis of CHO-K1 cell growth in a fixed bed bioreactor using magnetic resonance spectroscopy and imaging (1999)
    Springer
    Cytotechnology 30 (1999), S. 121-132 
    Details
    ISSN: 1573-0778
    Keywords: bioreactor ; cell volume ; imaging ; magnetic resonance
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Non-invasive magnetic resonance imaging and spectroscopy techniques have been used to monitor the growth and distribution of Chinese hamster ovary K1 cells growing in a fixed bed bioreactor composed of macroporous carriers. Diffusion-weighted 1H magnetic resonance spectroscopy was used to monitor the volume fraction of the bioreactor occupied by the cells and diffusion-weighted 1H magnetic resonance imaging was used to map cell distribution. The imaging measurements demonstrated that cell growth in the bioreactor was heterogeneous, with the highest cell densities being found at the surface of the carriers. The increase in the volume fraction occupied by the cells during cell growth showed a close correlation with bioreactor ATP content measured using 31P magnetic resonance spectroscopy. These magnetic resonance measurements, in conjunction with measurements of bioreactor glucose consumption, allowed estimation of the specific glucose consumption rate. This declined during the culture, in parallel with medium glucose concentration.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008039011960
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  • 6
    Electronic Resource
    Electronic Resource
    Use of cell cycle analysis to characterise growth and interferon-γ production in perfusion culture of CHO cells (1999)
    Springer
    Cytotechnology 30 (1999), S. 59-69 
    Details
    ISSN: 1573-0778
    Keywords: cell cycle ; CHO ; continuous culture ; flow cytometry ; perfusion culture ; spin-filter
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract The importance of cell cycle analysis in cell culture development has been widely recognised. Whether such analysis is useful in indicating future performance of high cell density culture is uncertain. Using flow cytometric approach to address this question, we utilised the fraction of cells in the S phase to control specific growth rate and productivity in spin filter perfusion cultures and found a significant increase in the accumulated interferon-γ over that obtained from the nutrient-based controlled fed culture. While a general decrease with time exists in both percentage of S phase cells and specific growth rate, a clear oscillatory behaviour of both parameters is found in perfusion cultures.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008055904642
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  • 7
    Electronic Resource
    Electronic Resource
    Degradative activities in a recombinant chinese hamster ovary cell culture (1996)
    Springer
    Cytotechnology 22 (1996), S. 43-52 
    Details
    ISSN: 1573-0778
    Keywords: recombinant CHO cells ; insulin degradative activity ; glycosidase ; bioreactor
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Two degradative activities were found in a recombinant Chinese hamster ovary cell culture. These activities became more dominant under high cell density and extended running time, as achieved in a semi-continous perfusion culture. The first, insulin degradative activity caused a growth upset in the 3rd cycle of the perfusion culture and shortened the length of the bioreactor process. The second activity, derived from the neutral pH stable sialidase, was found to affect the integrity of the carbohydrate structure of the recombinant protein, causing increase in heterogeneity in molecular weight and pI of the glycoforms. The most efficient way to overcome these problems may be the use of genetically altered ‘designer cells’ as the production cell line.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00353923
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  • 8
    Electronic Resource
    Electronic Resource
    Human diploid fibroblast growth on polystyrene microcarriers in aggregates (1996)
    Springer
    Cytotechnology 22 (1996), S. 111-117 
    Details
    ISSN: 1573-0778
    Keywords: aggregation ; bioreactor ; cell growth ; diploid fibroblasts ; microcarriers ; suspension
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Polystyrene microcarriers were prepared in four size ranges (53–63 μm, 90–125 μm, 150–180 μm and 300–355 μm) and examined for ability to support attachment and growth of human diploid fibroblasts. Cells attached rapidly to the microcarriers and there was a direct relationship between cell attachment and microcarrier aggregation. Phasecontrast and scanning electron microscopic studies revealed that while aggregation was extensive, most of the aggregate consisted of void volume. Cell growth studies demonstrated that human diploid fibroblasts proliferated well in microcarrier aggregates, reaching densities of 2.5–3×106 cells per 2 ml dish after 6 days from an inoculum of 0.5×106 cells per dish. When cells were added to the microcarriers at higher density (up to 5×106 cells per 2-ml culture), there was little net growth but the cells remained viable over a 7-day period. In contrast, cells died when plated under the same conditions in monolayer culture. When the microcarriers were used in suspension culture, rapid cell attachment and rapid microcarrier aggregation also occurred. In 100-ml suspension culture, a cell density of 0.7×106 cells per ml was reached after 7 days from an inoculum of 0.1×106 cells. Based on these data, we conclude that microcarrier aggregation is not detrimental to fibroblast growth. These data also indicate that small microcarriers (53–63 μm) (previously thought to be too small to support the growth of diploid fibroblasts) can support fibroblast growth and this occurs primarily because microcarriers in this size range efficiently form aggregates with the cells.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00353930
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  • 9
    Electronic Resource
    Electronic Resource
    Dialysis cultures with immobilized hybridoma cells for effective production of monoclonal antibodies (1997)
    Springer
    Cytotechnology 23 (1997), S. 39-45 
    Details
    ISSN: 1573-0778
    Keywords: fixed bed reactor ; immobilization ; dialysis technique ; hybridoma cells
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract An industrial scale reactor concept for continuous cultivation of immobilized animal cells (e.g. hybridoma cells) in a radial-flow fixed bed is presented, where low molecular weight metabolites are removed via dialysis membrane and high molecular products (e.g. monoclonal antibodies) are enriched. In a new “nutrient-split” feeding strategy concentrated medium is fed directly to the fixed bed unit, whereas a buffer solution is used as dialysis fluid. This feeding strategy was investigated in a laboratory scale reactor with hybridoma cells for production of monoclonal antibodies. A steady state monoclonal antibody concentration of 478 mg l-1 was reached, appr. 15 times more compared to the concentration reached in chemostat cultures with suspended cells. Glucose and glutamine were used up to 98%. The experiments were described successfully with a kinetic model for immobilized growing cells. Conclusions were drawn for scale-up and design of the large scale system. Abbreviations: cGlc – glucose concentration, mmol l-1; cGln – glutamine concentration, mmol l-1; cAmm – ammonia concentration, mmol l-1; cLac – lactate concentration, mmol l-1; cMAb – MAb concentration, mg l-1; D – dilution rate, d-1; Di – dilution rate in the inner chamber of the membrane dialysis reactor, d-1; D0 – dilution rate in the outer chamber of the membrane dialysis reactor, d-1; q*FB,Glc – volume specific glucose uptake rate related to the fixed bed volume, mmol lFB -1 h-1; q*FB,Gln – volume specific glutamine uptake rate related to the fixed bed volume, mmol lFB -1 h-1.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007911517505
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  • 10
    Electronic Resource
    Electronic Resource
    Hybridoma cell behaviour in continuous culture under hyperosmotic stress (1999)
    Springer
    Cytotechnology 29 (1999), S. 71-84 
    Details
    ISSN: 1573-0778
    Keywords: bioreactor ; continuous culture ; hybridoma cells ; hyperosmolality ; monoclonal antibody production ; non-producing subpopulation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract In this paper, we propose an alternative strategy to the ones proposed before (Oh et al., 1993; Øyaas et al., 1994a) to get real increases of global final antibody titer and production at hyperosmotic stress, by reducing the detrimental effect of such a stress on cell growth, and conserving the stimulating effect on antibody production. It consists of cultivating the cells in continuous culture and increasing the osmolality stepwise. In this way, the cells could progressively adapt to the higher osmolality at each step and antibody titers could be nearly doubled at 370 and 400 mOsm kg-1, compared to the standard osmolality of 335 mOsm kg-1. Surprisingly, the stimulation of antibody production was not confirmed for higher osmolalities, 425 and 450 mOsm kg- 1, despite the minor negative effect on cell growth. Intracellular IgG analysis by flow cytometry revealed at these osmolalities a significant population of non-producing cells. However, even when taking into account this non-producing population, a stimulating effect on antibody production could not be shown at these highest osmolalities. It seems to us that osmolality has a significant effect on the appearance of these non-producing cells, since they were not observed in continuous cultures at standard osmolality, of comparable duration and at an even higher dilution rate. The appearance of the non-producing cells coincides furthermore with modifications of the synthesised antibody, as shown by electrophoretic techniques. It is however not really clear if these two observations reflect actually the same phenomenon. Hyperosmolality affects the cell behaviour in continuous culture in multiple ways, independently of the growth rate, counting all at least partially for the observed stimulation of antibody production: acceleration of the amino acid, and in particular the glutamine metabolism, increase of the cell volume, increase of the intracellular pH and accumulation of cells in the G1 cell cycle phase.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008014909474
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  • 11
    Electronic Resource
    Electronic Resource
    Development of a fixed bed bioreactor for the expansion of human hematopoietic progenitor cells (1999)
    Springer
    Cytotechnology 30 (1999), S. 227-234 
    Details
    ISSN: 1573-0778
    Keywords: bioreactor ; cord blood ; expansion ; hematopoieticcells ; porous carrier ; stromal cells
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract The ex vivo expansion of hematopoietic progenitor cells is of great interest for a variety of clinical applications, e.g. bone marrow transplantation or gene therapy. Therefore it is of general interest to develop a culture system, able to mimic the in vivo hematopoesis, which is a prerequisite for long-term hematopoietic culture. Our approach was to modify a continuously perfused bioreactor for cultivation and expansion of human hematopoietic stem cells. Therefore we immobilized stromal cells (human primary stromal cells or the murine cell line M2-10B4) in porous glass carriers in a fixed bed reactor and cocultivated human hematopoietic progenitor cells for several weeks. After inoculation of mononuclear cells derived from umbilical cord blood or peripheral blood stem cells both adherent and non adherent cells were harvested and analyzed by flow cytometry and short-term colony assays. During cultivation there was a permanent production of progenitor cells and mature blood cells derived from the immobilized cells in the carriers. We could demonstrate the immobilization of hematopoietic progenitor cells of the myeloid system detectable in short-term colony assays. Additionally we could observe the expansion of very early progenitor cells (CFU-GEMM) up to 4.2-fold and later progenitor cells (CFU-GM and BFU-E) up to 7-fold and 1.8-fold, respectively.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008085932764
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  • 12
    Electronic Resource
    Electronic Resource
    Analysis of the use of fortified medium in continuous culture of mammalian cells (1999)
    Springer
    Cytotechnology 31 (1999), S. 243-254 
    Details
    ISSN: 1573-0778
    Keywords: continuous culture ; growth inhibition ; osmolality ; perfusion culture
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Continuous culture is frequently used in the cultivation of mammalian cells for the manufacturing of recombinant protein pharmaceuticals. In such operations a large volume of medium is turned over each day, especially in the case where cell recycle, or perfusion cultivation, is practiced. In principle, the volumetric throughput of medium can be reduced by using a more concentrated feed while maintaining the same nutrient provision rate. Overall, the medium components are divided into two categories: ‘consumable nutrients' and ‘unconsumable inorganic bulk salts’. In such fortified medium, the concentrations of consumable nutrients, but not bulk salts, are increased. With a stoichiometrically-balanced medium, the large amount of nutrients fed into the culture is largely consumed by cells to give rise to residual concentrations of these nutrients in their optimal range. However, unless care is taken to initiate the continuous culture, overshoot of nutrients may occur during the transient period. The high nutrient concentration during overshoot may be inhibitory by itself, or the resulting high osmolality may retard the growth. Using a mathematical model that incorporates the growth inhibitory effect of high osmolality we demonstrate such a potentially catastrophic effect of nutrient and osmolality overshoot by simulation. To avoid overshoot a controlled nutrient feeding scheme should be devised at the initiation of continuous culture.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008026613975
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  • 13
    Electronic Resource
    Electronic Resource
    Are we prepared for animal cell technology in the 21st century? (1995)
    Springer
    Cytotechnology 18 (1995), S. 3-8 
    Details
    ISSN: 1573-0778
    Keywords: bioreactor ; cellular therapies ; gene therapy ; therapeutic proteins
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Large scale animal cell culture for the production of complex therapeutic proteins has been a major success of the biotechnology industry. Today, approximately half of the $ 5 billion annual turnover of the biotechnology industry is based upon this technology, in many cases with reactors of more than 10 m3. As we look towards the 21 st century, however, we can see novel approaches to the production of therapeutic proteins, by means of gene and cellular therapies. These technologies present new engineering challenges to the animal cell technologist. Are we prepared to meet these challenges? The needs include: small-scale reactors for the preparation of autologous cell lines, methods for the production of viruses to be used as vectors in gene therapy, artificial organ and the processing of xenogenic cell lines and tissues for cellular implants in humans. More attention should be given to three-dimensional cell cultures. Mass transfer considerations need to be extended beyond just oxygen transfer, to include cellular communication in small systems; this is becoming increasingly important for the control and optimise growth and product formation. Apart from improvements of large-scale systems, substantial advantages could be gained by studying new methods for the production and delivery of therapeutic proteins, using small-scale cell culture systems. We should adapt teaching, regulatory, patent and clinical infrastructure to meet this challenge in a harmonious way.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00744313
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  • 14
    Electronic Resource
    Electronic Resource
    Oxygen gradients in animal-cell bioreactors (1995)
    Springer
    Cytotechnology 18 (1995), S. 27-34 
    Details
    ISSN: 1573-0778
    Keywords: Air lift reactor ; bubble column ; bioreactor ; oxygen gradients ; scale-up ; stirred vessel
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract An estimation is made of oxygen gradients in animal-cell bioreactors, using straightforward engineering calculations. Three types of bioreactor are considered: stirred vessel, bubble column and air lift, of sizes between 0.01 and 10 m3. First, the gradient is estimated in the stagnant layer surrounding a cell (15 μm), a microcarrier (185 μm) with 300 cells attached to it, a macroporous support (1.25 mm) containing 185,00 cells and one (6 mm) containing 4.25 million cells. It is assumed that oxygen consumption is 10−16 mole O2·cell−1·s−1, while mass transfer coefficients are obtained from Sherwood relations. Circulation and liquid-retention times of the bioreactors are compared with the oxygen-exhaust times of suspensions with 1012, 1013 and 1014 cells/m3 to estimate if oxygen gradients are likely to exist in the bulk-liquid phase. Finally, the gradient in the liquid film surrounding air bubbles is estimated using k l A-values obtained from empirical correlations. It is clear from all these estimations that in many situations severe gradients can be expected. The question remains, however, whether gradients should be avoided as much as possible, or may be tolerated to a certain extent or even created on purpose because of possible beneficial effects.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00744317
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  • 15
    Electronic Resource
    Electronic Resource
    Batch control system vaccines: BCSV (1995)
    Springer
    Cytotechnology 18 (1995), S. 57-66 
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    ISSN: 1573-0778
    Keywords: Automation ; bioreactor ; optimisation ; process control ; software sensors ; validation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract The Batch Control System for Vaccines (BCSV), a new Man Machine Interface (MMI) for the control of cultivations in bioreactors, was developed according to SP-88. SP-88 is the ISA standard for Batch Control Systems. Among others, SP-88 supplied the concept of recipes, which organize and specify the monitoring and control requirements for manufacturing. Process optimisation and compliance to GMP rules and regulations were the main objectives for this development. The most important features of the BCSV interface include: - implementation at production, pilot and R & D scale to assure easy transfer of knowledge and experience at the various stage of process development; - independency of underlying hardware to ensure similar “look and feel” for different pieces of equipment; - in-house development and maintenance of recipes to have maximum control over applications; - interactive communication between operator and BCSV during recipe execution. GMP compliance was assured not only by considering governing sets of GMP regulations, but also by taking up the interface in a overall Information & Automation strategy and by setting up a QA strategy for the entire life cycle of the system.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00744320
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  • 16
    Electronic Resource
    Electronic Resource
    Ex vivo expansion of primitive hematopoietic cells for cellular therapies: An overview (1995)
    Springer
    Cytotechnology 18 (1995), S. 133-146 
    Details
    ISSN: 1573-0778
    Keywords: ex vivo expansion ; hematopoietic culture ; bioreactor ; clinical therapies ; cytokines ; stroma
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Sources of hematopoietic cells for bone marrow transplantation are limited by the supply of compatible donors, the possibility of viral infection, and autologous (patient) marrow that is depleted from prior chemo- or radiotherapy or has cancerous involvement. Anex vivo system to amplify hematopoietic progenitor cells could increase the number of patients eligible for autologous transplant, allow use of cord blood hematopoietic cells to repopulate an adult, reduce the amount of bone marrow and/or mobilized peripheral blood stem and progenitor cells required for transplantation, and reduce the time to white cell and platelet engraftment. The cloning of hematopoietic growth factors and the identification of appropriate conditions has enabled the development of successfulex vivo hematopoietic cell cultures. Purification systems based on the CD34 marker (which is expressed by the most primitive hematopoietic cells) have proven an essential tool for research and clinical applications. Present methods for hematopoietic cultures (HC) on stromal (i.e. accessory cells that support hematopoiesis) layers in flasks lack a well-controlled growth environment. Several bioreactor configurations have been investigated, and a first generation of reactors and cultures has reached the clinical trial stage. Our research suggests that perfusion conditions improve substantially the performance of hematopoietic reactors. We have designed and tested a perfusion bioreactor system which is suitable for the culture of non-adherent cells (without stromal cells) and readily scaleable for clinical therapies. Eliminating the stromal layer eliminates the need for a stromal cell donor, reduces culture time, and simplifies the culture system. In addition, we have compared the expansion characteristics of both mononuclear and CD34+ cells, since the latter are frequently assumed to give a superior performance for likely transplantation therapies.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00744329
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  • 17
    Electronic Resource
    Electronic Resource
    Changes in monoclonal antibody productivity of recombinant BHK cells immobilized in collagen gel particles (1997)
    Springer
    Cytotechnology 23 (1997), S. 5-12 
    Details
    ISSN: 1573-0778
    Keywords: monoclonal antibody ; immobilization ; collagen gel ; BHK ; productivity ; recombinant ; high density culture
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Animal cell perfusion high density culture is often adopted for the production of biologicals in industry. In high density culture sometimes the productivity of biologicals has been found to be enhanced. Especially in immobilized animal cell culture, significant increase in the productivity has been reported. We have found that the specific monoclonal antibody (MAb) productivity of an immobilized hybridoma cell is enhanced more than double. Several examples of enhancing productivities have been also shown by collagen immobilized cells. Immobilized cells involve some different points from non-immobilized cells in high density culture: In immobilized culture, some cells are contacted together, resulting in locally much higher cell concentration more than 108 cells/ml. Information originating from a cell can be easily transduced to the others in immobilized culture because the distance between cells is much nearer. Here we have performed collagen gel immobilized culture of recombinant BHK cells which produce a human IgG monoclonal antibody in a protein-free medium for more than three months. In this high density culture a stabilized monoclonal antibody production was found with around 8 times higher specific monoclonal antibody productivity compared with that in a batch serum containing culture. No higher MAb productivity was observed using a conditioned medium which was obtained from the high density culture, indicating that no components secreted from the immobilized cells work for enhancing monoclonal antibody production. The MAb productivity by the non-immobilized cells obtained by dissolving collagen using a collagenase gradually decreased and returned to the original level in the batch culture using a fresh medium. This suggests that the direct contact of the cells or a very close distance between the cells has something to do with the enhancement of the MAb productivity, and the higher productivity is kept for a while in each cell after they are drawn apart.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007959400666
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  • 18
    Electronic Resource
    Electronic Resource
    Comparison of manufacturing techniques for adenovirus production (1999)
    Springer
    Cytotechnology 30 (1999), S. 169-172 
    Details
    ISSN: 1573-0778
    Keywords: adenovirus ; bioreactor ; microcarriers ; serum-free medium ; 293 cells
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract We have compared three different production methods, which may be suitable for the large scale production of adenovirus vectors for human clinical trials. The procedures compared 293 cells adapted to suspension growth in serum-free medium in a stirred tank bioreactor, 293 cells on microcarriers in serum-containing medium in a stirred tank bioreactor, and 293 cells grown in standard tissue culture plasticware. With a given virus, yields varied between 2000 and 10,000 infectious units/cell. The stirred tank bioreactor routinely produced between 4000 and 7000 infectious units/cell when 293 cells were grown on microcarriers. The 293 cells adapted to suspension growth in serum-free medium in the same stirred tank bioreactor yielded between 2000 and 7000 infectious units/cell. Yields obtained from standard tissue culture plasticware were up to 10,000 infectious units/cell. Cell culture conditions were monitored for glucose consumption, lactate production, and ammonia accumulation. Glucose consumption and lactate accumulation correlated well with the cell growth parameters. Ammonia production does not appear to be significant. Based on virus yields, ease of operation and linear scalability, large-scale adenovirus production seems feasible using 293 cells (adapted to suspension/serum free medium or on microcarriers in serum containing medium) in a stirred tank bioreactor.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008040221630
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  • 19
    Electronic Resource
    Electronic Resource
    Xanthomonas campestris as a host for the production of recombinantPseudomonas aeruginosa lipase (1996)
    Springer
    Journal of industrial microbiology and biotechnology 16 (1996), S. 22-28 
    Details
    ISSN: 1476-5535
    Keywords: lipase ; recombinantXanthomonas ; fed-batch ; bioreactor
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Recombinant plasmid pBP13, which expresses the alkaline lipase fromPseudomonas aeruginosa IGB83 under thetac promoter was transferred toXanthomonas campestris pvcampestris IBT148. Different fermentation conditions were tested for lipase productivity by strain IBT148 carrying plasmid pBP13, and a fermentation process was established in an instrumented bioreactor, where lipase production was increased more than 12-fold with respect to the initial culture conditions in shake flasks. Xanthan gum stabilized the activity of the alkaline lipase.
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    Glucose oxidase and catalase activities ofPenicillium variabile P16 immobilized in polyurethane sponge (1996)
    Springer
    Journal of industrial microbiology and biotechnology 17 (1996), S. 15-19 
    Details
    ISSN: 1476-5535
    Keywords: glucose oxidase ; catalase ; Penicillium variabile ; immobilization ; polyurethane sponge
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Conidia ofPenicillium variabile P16 were immobilized in polyurethane sponge and used in repeated-batch processes in a fluidized-bed reactor. Optimal conditions for production of glucose oxidase and catalase were: inoculum size, 10%; glucose concentration, 80 g L−1; Ca-carbonate concentration, 15 g L−1; temperature, 28°C and aeration rate, 4 VV−1 min−1. In an extended repeated-batch process, glucose oxidase activity was highest after the fourth batch and catalase activity was highest after the fifth batch. Scanning electron microscopy showed that the fungus grew only in the interior of carrier particles.
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    URL: http://dx.doi.org/10.1007/BF01570142
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    Ethanol production by immobilized whole cells ofZymomonas mobilis in a continuous flow expanded bed bioreactor and a continuous flow stirred tank bioreactor (1996)
    Springer
    Journal of industrial microbiology and biotechnology 17 (1996), S. 11-14 
    Details
    ISSN: 1476-5535
    Keywords: continuous flow reactor ; ethanol ; expanded bed reactor ; immobilization ; Zymomonas mobilis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Continuous ethanol fermentation by immobilized whole cells ofZymomonas mobilis was investigated in an expanded bed bioreactor and in a continuous stirred tank reactor at glucose concentrations of 100, 150 and 200 g L−1. The effect of different dilution rates on ethanol production by immobilized whole cells ofZymomonas mobilis was studied in both reactors. The maximum ethanol productivity attained was 21 g L−1 h−1 at a dilution rate of 0.36 h−1 with 150 g glucose L−1 in the continuous expanded bed bioreactor. The conversion of glucose to ethanol was independent of the glucose concentration in both reactors.
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    URL: http://dx.doi.org/10.1007/BF01570141
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    Biotransformation of codeine to morphine in freely suspended cells and immobilized cultures of Spirulina platensis (1999)
    Springer
    World journal of microbiology and biotechnology 15 (1999), S. 465-469 
    Details
    ISSN: 1573-0972
    Keywords: Biotransformation ; codeine ; immobilization ; morphine ; Spirulina platensis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Both freely suspended cells and immobilized cultures of Spirulina platensis, a blue-green alga, biotransformed exogenously fed codeine, an opium alkaloid, to morphine. The external addition of codeine to the culture medium did not affect the growth of S. platensis. Immobilization of Spirulina in a calcium alginate gel matrix was optimized by using 2% (w/v) sodium alginate and reducing the concentration of nutrients of Zarrouk's medium, which caused destabilization of the calcium alginate gel. The accumulation of morphine increased gradually and reached maxima of 330 μg 100 ml−1 culture at 105 h in freely suspended and 351 μg 100 ml−1 at 96 h in immobilized Spirulina cultures. Accumulation of morphine was detected only in the medium, whereas cells did not show accumulation. The immobilized Spirulina cultures showed marginally higher conversion of codeine to morphine over freely suspended cultures.
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    Biodegradation of olive oil mill wastewaterby Coriolus versicolor and Funalia trogii:effects of agitation, initial COD concentration, inoculum size and immobilization (1997)
    Springer
    World journal of microbiology and biotechnology 14 (1997), S. 37-42 
    Details
    ISSN: 1573-0972
    Keywords: Biodegradation ; immobilization ; laccase ; olive oil mill wastewater ; white rot fungi
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract The biodegradation of olive oil mill wastewater (OOMW) by Coriolus versicolor and Funalia trogii was investigated. Initial COD concentration, agitation and inoculum size were all found to be significant for biodegradation. Adding glucose, sulphate or nitrogen had no effect on biodegradation. During growth in optimum conditions, C.versicolor removed approximately 63% COD, 90% phenol and 65% colour within 6 days and F. trogii removed approximately 70% COD, 93% phenol and 81% colour of the OOMW used. The fungi also excreted large amounts of extracellular laccase into the medium. High biodegradation yields were also obtained by fungi immobilized in calcium alginate gels.
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    The use of monochloroacetic acid for improved ethanol production by immobilized Saccharomyces cerevisiae (1997)
    Springer
    World journal of microbiology and biotechnology 14 (1997), S. 107-111 
    Details
    ISSN: 1573-0972
    Keywords: Glutaraldehyde ; immobilization ; monochloroacetic acid ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
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    URL: http://dx.doi.org/10.1023/A:1008888820176
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    Glucoamylase production in continuous cultures of Aspergillus niger with special emphasis on growth parameters (1997)
    Springer
    World journal of microbiology and biotechnology 14 (1997), S. 113-118 
    Details
    ISSN: 1573-0972
    Keywords: Aspergillus ; continuous culture ; glucoamylase ; growth ; fungi ; nitrogen
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Maltose-limited continuous culture of Aspergillus niger was carried out with potassium nitrate to investigate growth and glucoamylase formation characteristics. Glucoamylase production was dependent on the specific growth rate. The maximal amount of glucoamylase (units/l and U/g dry weight) was obtained at μ=0.08h−1, and the maximum specific rate of production (units/g/dry weight per hour) was at μ=0.2h−1. The maintenance coefficients (ms and mATP) were higher than for some other fungi. Maximal growth yields on substrate, oxygen and ATP (Yxsm, YxO2m and Yxam) were very efficient (high) and the value of Yxam, which cannot exceed the theoretical maximal value, is obtained when a P/O ratio of 1:1 is assumed. This indicates that biomass formation is energetically inexpensive and most of the expended energy has to be invested in the process of glucoamylase excretion.
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    URL: http://dx.doi.org/10.1023/A:1008840904246
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    Kinetic characterization in batch and continuous culture of Escherichia coli mutants affected in phosphoenolpyruvate metabolism: differences in acetic acid production (1999)
    Springer
    World journal of microbiology and biotechnology 15 (1999), S. 587-592 
    Details
    ISSN: 1573-0972
    Keywords: Acetic acid production ; carbon metabolism ; continuous culture ; Escherichia coli ; metabolic engineering
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract The growth kinetics of an Escherichia coli wild type strain and two derivative mutants were examined in batch cultures and in glucose-limited chemostats. One mutant (PB12) had an inactive phosphotranferase transport system and the other (PB25) had interrupted pykA and pykF genes that code for the two pyruvate kinase isoenzymes. In both batch and continuous culture, important differences in acetic acid accumulation and other metabolic activities were found. Compared to the wild type strain, we observed a reduction in acetic acid accumulation of 25 and 80% in PB25 and PB12 strains respectively, in batch culture. Continuous culture experiments revealed that compared to the other two strains, PB25 accumulated less acetic acid as a function of dilution rate. In continuous cultures, oxidoreductase metabolic activities were substantially affected in the two mutant strains. These changes in turn were reflected in different levels of biomass and CO2 production, and in oxygen consumption.
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    Production, extraction and purificationof violacein: an antibiotic pigment producedby Chromobacterium violaceum (1998)
    Springer
    World journal of microbiology and biotechnology 14 (1998), S. 685-688 
    Details
    ISSN: 1573-0972
    Keywords: Antibiotic ; bioreactor ; Chromobacterium violaceum ; pigment ; violacein
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract A procedure for the production, extraction, and purification of violacein was developed using Chromobacterium violaceum (CCT 3496) cultivated on cotton, in modified 1 litre Roux bottles. A surface tray bioreactor was built to perform these experiments. Violacein was extracted with commercial ethanol, and purified by filtration, Soxhlet extraction, crystallization and high performance liquid chromatography. The violacein was analysed and identified by proton and carbon-13 NMR spectroscopies, thermogravimetric analysis, mass spectrometry, UV-VIS spectroscopy and infrared spectroscopy. It was concluded that the product was highly purified violacein.
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    High-density perfusion culture of insect cells with a BioSep ultrasonic filter (1998)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 59 (1998), S. 351-359 
    Details
    ISSN: 0006-3592
    Keywords: bioreactor ; high density ; insect cells ; perfusion ; Sf9 ; ultrasonic filter ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The baculovirus/insect cell expression system has provided a vital tool to produce a high level of active proteins for many applications. We have developed a very high-density insect cell perfusion process with an ultrasonic filter as a cell retention device. The separation efficiency of the filter was studied under various operating conditions. A cell density of over 30 million cells/mL was achieved in a controlled perfusion bioreactor and cell viability remained greater than 90%. Sf9 cells from a high-density culture and a spinner culture were infected with two recombinant baculoviruses expressing genes for the production of human chitinase and monocyte-colony inhibition factor. The protein yield on a cell basis from infecting high-density Sf9 cells was the same as or higher than that from the spinner Sf9 culture. Virus production from the high-density culture was similar to that from the spinner culture. The results show that the ultrasonic filter did not affect insect cells' ability to support protein expression and virus production following infection with baculovirus. The potential applications of the high-density perfusion culture for large-scale protein expression from Sf9 cells are also highlighted. © 1998 John Wiley & Sons, Inc. Biotechnol Bioeng 59:351-359, 1998.
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    On-line monitoring of respiration in recombinant-baculovirus infected and uninfected insect cell bioreactor cultures (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 50 (1996), S. 36-48 
    Details
    ISSN: 0006-3592
    Keywords: insect cell culture ; Sf-9 cells ; respiration ; bioreactor ; on-line monitoring ; baculovirus expression vector system ; recombinant proteins ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Respiration rates in Spodoptera frugiperda (Sf-9) cell bioreactor cultures were successfully measured on-line using two methods: The O2 uptake rate (OUR) was determined using gas phase pO2 values imposed by a dissolved oxygen controller and the CO2 evolution rate (CER) was measured using an infrared detector. The measurement methods were accurate, reliable, and relatively inexpensive. The CER was routinely determined in bioreactor cultures used for the production of several recombinant proteins. Simple linear relationships between viable cell densities and both OUR and CER in exponentially growing cultures were used to predict viable cell density. Respiration measurements were also used to follow the progress of baculoviral infections in Sf-9 cultures. Infection led to increases in volumetric and per-cell respiration rates. The relationships between respiration and several other culture parameters, including viable cell density, cell protein, cell volume, glucose consumption, lactate production, viral titer, and recombinant β-galactosidase accumulation, were examined. The extent of the increase in CER following infection and the time postinfection at which maximum CER was attained were negatively correlated with the multiplicity of infection (MOI) at multiplicities below the level required to infect all the cells in a culture. Delays in the respiration peak related to the MOI employed were correlated with delays in the peak in recombinant protein accumulation. DO levels in the range 5-100% did not exert any major effects on viable cell densities, CER, or product titer in cultures infected with a baculovirus expressing recombinant β-galactosidase. © 1996 John Wiley & Sons, Inc.
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    Kinetic studies of fusarium solani pisi cutinase used in a gas/solid system: Transesterification and hydrolysis reactions (1997)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 56 (1997), S. 1-8 
    Details
    ISSN: 0006-3592
    Keywords: transesterification ; hydrolysis ; water activity ; cutinase ; gas ; bioreactor ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Fusarium solani cutinase supported onto Chromosorb P was used to catalyze transesterification (alcoholysis) and hydrolysis on short volatile alcohols and esters in a continuous gas/solid bioreactor. In this system, a solid phase composed of a packed enzymatic preparation was continuously percolated with carrier gas which fed substrates and removed reaction products simultaneously. A kinetic study was performed under differential operating conditions in order to get initial reaction rates. The effect of the hydration state of the biocatalyst on the kinetics was studied for 3 conditions of hydration (aw = 0.2, aw = 0.4 and aw = 0.6), the alcoholysis of propionic acid methyl ester with n-propanol, and for 5 hydration levels (from aw = 0.2 to aw = 0.6) for the hydrolysis of propionic acid methyl, ethyl or propyl esters. F. solani cutinase was found to have an unusual kinetic behavior. A sigmoid relationship between the rate of transesterification and the activity of methyl propionate was observed, suggesting some form of cooperative activation of the enzyme by one of its substrate. For the hydrolysis of short volatile propionic acid alkyl esters, threshold effects on the reaction rate, highly depending on the water activity and the substrate polarity, are reported. © 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 56: 1-8, 1997.
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    Perfusion bioreactors for the production of recombinant proteins in insect cells (1996)
    Springer
    Cytotechnology 20 (1996), S. 191-198 
    Details
    ISSN: 1573-0778
    Keywords: insect cell culture ; perfusion culture ; membrane perfusion ; crossflow microfiltration ; baculovirus ; bioreactor ; fluidized bed ; packed bed ; recombinant protein production
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Conclusion High density perfusion culture of insect cells for the production of recombinant proteins has proved to be an attractive alternative to batch and fed-batch processes. A comparison of the different production processes is summarized in Table 3. Internal membrane perfusion has a limited scale-up potential but appears to the method of choice in smaller lab-scale production systems. External membrane perfusion results in increased shear stress generated by pumping of cells and passing through microfiltration modules at high velocity. However, using optimized perfusion strategies this shear stress can be minimized such that it is tolerated by the cells. In these cases, perfusion culture has proven to be superior to batch production with respect to product yields and cell specific productivity. Although insect cells could be successfully cultivated by immobilization and perfusion in stationary bed bioreactors, this method has not yet been used in continuous processes. In fluidized bed bioreactors with continuous medium exchange cells showed reduced growth and protein production rates. For the cultivation of insect cells in batch and fedbatch processes numerous efforts have been made to optimize the culture medium in order to allow growth and production at higher cell densities. These improved media could be used in combination with a perfusion process, thus allowing substantially increased cell densities without raising the medium exchange rate. However, sufficient oxygen supply has to be guaranteed during fermentation in order to ensure optimal productivity.
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    Growth Stimulation of Tumor-Specific Cytotoxic T Lymphocytes on Concanavalin A-Immobilized Carrier Beads (1998)
    Springer
    Cytotechnology 26 (1998), S. 13-21 
    Details
    ISSN: 1573-0778
    Keywords: concanavalin A ; cytotoxic T lymphocytes ; immobilization ; interleukin
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Human tumor-specific CD4+ cytotoxic T lymphocytes (CTL) were generated against duodenum papilloma cell line TGBC18TKB from HLA type-matched peripheral blood mononuclear cells. Concanavalin A (Con A) immobilized on carrier beads stimulated growth of the CTL in a long-term culture without repeated antigen stimulation, while soluble Con A induced death of the CTL. The CTL exhibited the target-specific cytotoxicity in a more potent manner than those before the long-term culture in the presence of the immobilized Con A. Enhanced expression of the adhesion molecule, CD11b, was observed on the CTL. These results suggest that immobilized Con A will be useful for continuous growth stimulation and large scale expansion of CTL without tumor antigen.
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    Development of a versatile computer integrated control system for bioprocess controls (1998)
    Springer
    Cytotechnology 26 (1998), S. 227-236 
    Details
    ISSN: 1573-0778
    Keywords: bioreactor ; computer control ; data acquisition ; glucose control
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract A general approach is described for the implementation of a networked multi-unit computer integrated control system. The use of data acquisition hardware and graphical programming tools alleviates tedious programming and maintains potency and flexibility. One application of the control system, the control of a mammalian cell perfusion culture based on a key nutrient glucose concentration, was demonstrated. The control system offers customized user interface for all process control parameters and allows the flexibility for continued improvement and implementation of new tailored functions. The temperature, pH, dissolved oxygen and glucose level were accurately controlled.
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    Effects of CO2 and osmolality on hybridoma cells: growth, metabolism and monoclonal antibody production (1998)
    Springer
    Cytotechnology 28 (1998), S. 213-227 
    Details
    ISSN: 1573-0778
    Keywords: antibody production ; carbon dioxide ; cell metabolism ; continuous culture ; inhibition
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract CO2 partial pressure (pCO2) in industrial cell culture reactors may reach 150–200 mm Hg, which can significantly inhibit cell growth and recombinant protein production. Due to equilibrium with bicarbonate, increased pCO2 at constant pH results in a proportional increase in osmolality. Hybridoma AB2-143.2 cell growth rate decreased with increasing pCO2 in well-plate culture, with a 45% decrease at 195 mm Hg with partial osmolality compensation (to 361 mOsm kg- 1). Inhibition was more extensive without osmolality compensation, with a 63% decrease in growth rate at 195 mm Hg and 415 mOsm kg-1. Also, the hybridoma death rate increased with increasing pCO2, with 31- and 64-fold increases at 250 mm Hg pCO2 for 401 and 469 mOsm kg- 1, respectively. The specific glucose consumption and lactate production rates were 40–50% lower at 140 mm Hg pCO2. However, there was little further inhibition of glycolysis at higher pCO2. The specific antibody production rate was not significantly affected by pCO2 or osmolality within the range tested. Hybridomas were also exposed to elevated pCO2 in continuous culture. The viable cell density decreased by 25–40% at 140 mm Hg. In contrast to the well-plate cultures, the death rate was lower at the new steady state at 140 mm Hg. This was probably due to higher residual nutrient and lower byproduct levels at the lower cell density (at the same dilution rate), and was associated with increased cell-specific glucose and oxygen uptake. Thus, the apparent effects of pCO2 may vary with the culture system.
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    A microcarrier-based cell culture process for the production of a bovine respiratory syncytial virus vaccine (1999)
    Springer
    Cytotechnology 29 (1999), S. 135-149 
    Details
    ISSN: 1573-0778
    Keywords: anchorage dependent ; animal cell ; bioreactor ; Cultispher S ; Cytodex 3 ; microcarrier ; respiratory syncytial virus ; vaccine
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Veterinary viral vaccines generally comprise either attenuated or chemically inactivated viruses which have been propagated on mammalian cell substrates or specific pathogen free (SPF) eggs. New generation vaccines include chemically inactivated virally-infected whole cell vaccines. The NM57 cell line is a bovine nasal turbinate persistently infected (non-lytic infection) with a strain of the respiratory syncytial virus (RSV). The potential of microcarrier technology for the cultivation in bioreactors of this anchorage dependent cell line for RSV vaccine production has been investigated. Both Cytodex 3 and Cultispher S microcarriers proved most suitable from a selection of microcarriers as growth substrates for this NM57 cell line. Maximum cell densities of 4.12×105 cells ml-1and 5.52×105 cells ml-1 respectively were obtained using Cytodex 3 (3 g l-1) and and Cultispher S (1 g l-1) in 5 l bioreactor cultures. The fact that cell growth was less sensitive to agitation rate when cultured on Cultispher S microcarriers, and that cells were efficiently harvested from this microcarrier by an enzymatic method, suggested Cultispher S is suitable for further evaluation at larger bioreactor scales (〉5 l) than that described here.
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    Production of recombinant h-AT III with mammalian cell cultures using capillary electrophoresis for product monitoring (1996)
    Springer
    Cytotechnology 21 (1996), S. 205-215 
    Details
    ISSN: 1573-0778
    Keywords: antithrombin III ; mammalian cell culture ; continuous culture ; capillary electrophoresis ; product monitoring ; recombinant protein ; temperature influence
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract The importance of mammalian cell cultures for biotechnological production processes is steadily increasing, despite the high demands of these organisms on their culture conditions. Efforts towards a more efficient bioprocess generally concentrate on maximizing the culture's life time, the cell number, and the product concentration. Here recombinant BHK 21 c13 cells are used to produce rh-AT III, an anticoagulant of high therapeutic value. The influence of the process mode (batch, repeated batch, continuous perfusion) and the process temperature (30°C vs. 37°C) on the above mentioned parameters is investigated. It is possible to increase the length of the culture from 140 h (batch) to more than 500 h (continuous perfusion culture), while concomitantly increasing the cell density from 0.72 106/ml (batch) to 2.27 106/ml (repeated batch) and 2.87 106/ml (continuous perfusion culture). The accumulation of toxic metabolites, such as lactate, can be curtailed by reducing the bioreactor temperature from 37°C to 30°C during the later part of the exponential growth phase. Fast and reliable product monitoring became essential during process optimization. Capillary zone electrophoresis (CZE) in uncoated fused silica capillaries was studied for that purpose and compared to the standard ELISA. Under optimized conditions an AT III quantification could be done within 2 min with CZE. The detection limit was 5 μg/ml. A relative standard deviation of less than 0.9% was calculated. The detection limit could be lowered by one order of magnitude by using a two dimensional system, where an liquid chromatographic (LC) system is coupled to the CZE. Concomitantly the resolution is improved. The two-dimensional analysis required 5 min. Membrane adsorbers (MA) were used as stationary phase in the LC-system, to allow the application of high flow rates (5–10 ml/min). The correlation between the LC-CZE analysis and the standard AT III-ELISA was excellent, with r2: 0.965. Using the assay for at line product monitoring, it is shown, that the process temperature is of no consequence for the productivity whereas the process mode strongly influences this parameter.
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    Foreign protein expression from S phase specific promoters in continuous cultures of recombinant CHO cells (1996)
    Springer
    Cytotechnology 22 (1996), S. 179-184 
    Details
    ISSN: 1573-0778
    Keywords: cell cycle ; CHO cells ; continuous culture ; SV40 promoter
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Foreign protein expression from the commonly used SV40 promoter has been found to be primarily during the S-phase of the cell cycle. Simple mathematical models with this cell cycle phase dependent expression of foreign protein suggest that the specific production rate will be proportional to the cell growth rate, which is particularly disadvantageous in high cell density fed-batch or perfusion bioreactors. In this study we investigate this predicted relationship between the production rate and growth rate by culturing recombinant CHO cells in a continuous suspension bioreactor. One CHO cell line, GS-26, has been stably transfected with the plasmid pSVgal, which contains the E. coli lac Z gene under the control of the SV40 promoter. This GS-26 cell line was grown in suspension cultures over a range of specific growth rates in batch and continuous modes. The intracellular β-galactosidase activity was assayed using a standard spectrophotometric method after breaking the cells open and releasing the enzyme. A strong growth associated relationship is found between the intracellular β-galactosidase content and the specific growth rate in batch and continuous cultures, as predicted.
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    Disposable bioreactor for cell culture using wave-induced agitation (1999)
    Springer
    Cytotechnology 30 (1999), S. 149-158 
    Details
    ISSN: 1573-0778
    Keywords: bioreactor ; cell culture ; disposable ; wave agitation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract This work describes a novel bioreactor system for the cultivation of animal, insect, and plant cells using wave agitation induced by a rocking motion. This agitation system provides good nutrient distribution, off-bottom suspension, and excellent oxygen transfer without damaging fluid shear or gas bubbles. Unlike other cell culture systems, such as spinners, hollow-fiber bioreactors, and roller bottles, scale-up is simple, and has been demonstrated up to 100 L of culture volume. The bioreactor is disposable, and therefore requires no cleaning or sterilization. Additions and sampling are possible without the need for a laminar flow cabinet. The unit can be placed in an incubator requiring minimal instrumentation. These features dramatically lower the purchase cost, and operating expenses of this laboratory/pilot scale cell cultivation system. Results are presented for various model systems: 1) recombinant NS0 cells in suspension; 2) adenovirus production using human 293 cells in suspension; 3) Sf9 insect cell/baculovirus system; and 4) human 293 cells on microcarrier. These examples show the general suitability of the system for cells in suspension, anchorage-dependent culture, and virus production in research and GMP applications.
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    Engineering Chinese hamster ovary (CHO) cells to achieve an inverse growth – associated production of a foreign protein, β-galactosidase (1998)
    Springer
    Cytotechnology 28 (1998), S. 73-80 
    Details
    ISSN: 1573-0778
    Keywords: adenovirus major late promoter ; β-galactosidase ; Chinese hamster ovary cells ; continuous culture ; G1 phase expression ; inverse-growth associated production
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Protein synthesis in mammalian cells can be observed in two strikingly different patterns: 1) production of monoclonal antibodies in hybridoma cultures is typically inverse growth associated and 2) production of most therapeutic glycoproteins in recombinant mammalian cell cultures is found to be growth associated. Production of monoclonal antibodies has been easily maximized by culturing hybridoma cells at very low growth rates in high cell density fed- batch or perfusion bioreactors. Applying the same bioreactor techniques to recombinant mammalian cell cultures results in drastically reduced production rates due to their growth associated production kinetics. Optimization of such growth associated production requires high cell growth conditions, such as in repeated batch cultures or chemostat cultures with attendant excess biomass synthesis. Our recent research has demonstrated that this growth associated production in recombinant Chinese hamster ovary (CHO) cells is related to the S (DNA synthesis)-phase specific production due to the SV40 early promoter commonly used for driving the foreign gene expression. Using the stably transfected CHO cell lines synthesizing an intracellular reporter protein under the control of SV40 early promoter, we have recently demonstrated in batch and continuous cultures that the product synthesis is growth associated. We have now replaced this S-phase specific promoter in new expression vectors with the adenovirus major late promoter which was found to be active primarily in the G1-phase and is expected to yield the desirable inverse growth associated production behavior. Our results in repeated batch cultures show that the protein synthesis kinetics in this resulting CHO cell line is indeed inverse growth associated. Results from continuous and high cell density perfusion culture experiments also indicate a strong inverse growth associated protein synthesis. The bioreactor optimization with this desirable inverse growth associated production behavior would be much simpler than bioreactor operation for cells with growth associated production.
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    URL: http://dx.doi.org/10.1023/A:1008069312131
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    Production, purification and immobilization of glucose isomerase from Streptomyces olivochromogenes PTCC 1547 (1997)
    Springer
    World journal of microbiology and biotechnology 13 (1997), S. 597-598 
    Details
    ISSN: 1573-0972
    Keywords: Glucose isomerase ; immobilization ; production ; purification ; Streptomyces olivochromogenes PTCC 1457
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Production of glucose isomerase from Streptomyces olivochromogenes PTCC 1457 was followed by its purification and immobilization. Different immobilization methods including the use of a hydrophobic support were investigated.
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    URL: http://dx.doi.org/10.1023/A:1018590031137
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    Effect of surfactants on the production of ergot-alkaloids by immobilized mycelia of Claviceps paspali (1998)
    Springer
    World journal of microbiology and biotechnology 14 (1998), S. 447-450 
    Details
    ISSN: 1573-0972
    Keywords: Claviceps ; ergot alkaloids ; immobilization ; surfactant
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract In a semicontinuous process immobilized Claviceps paspali mycelia produced alkaloids over a period of 60 days (six reincubations). By addition of the surfactant Pluronik, a polyethoxypolypropoxy polymer, a considerable increase in alkaloid biosynthesis occurred. The maximum product concentration achieved was 8.35gl-1, and the overall productivity was 5.80 mgl-1 h-1, which is half the productivity of the batch process. Maximum process productivity for a single reincubation (12.3 mg l-1 h-1) was almost equal to the batch process productivity.
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    URL: http://dx.doi.org/10.1023/A:1008837916873
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    Performance and efficiency of a yeast biofilter for the treatment of a Nigerian fertilizer plant effluent (1999)
    Springer
    World journal of microbiology and biotechnology 15 (1999), S. 515-516 
    Details
    ISSN: 1573-0972
    Keywords: Biofilter ; biodegradation ; effluent ; fertilizer ; immobilization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract A biofilter composed of yeasts and cassava peel was used to detoxify fertilizer plant effluent. The biological oxygen demand was reduced on treatment from a range of 1200–1400 mg/l to a range 135–404 mg/l. The ammonia-nitrogen (NH3–N) and nitrate-nitrogen (NO3–N) were reduced after treatment from 1000 to 10 mg/l and from 100 to 17.6 mg/l, respectively. The biofilter is simple and easy to handle with high efficiency of 98%.
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    Short communication: Ethanol production from cellulose at 45°C using a batch-fed system containing alginate-immobilized Kluyveromyces marxianus IMB3 (1996)
    Springer
    World journal of microbiology and biotechnology 12 (1996), S. 103-104 
    Details
    ISSN: 1573-0972
    Keywords: Alginate ; cellulase ; cellulose ; ethanol ; immobilization ; Kluyveromyces marxianus
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract The thermotolerant yeast, Kluyveromyces marxianus IMB3, was grown in batch culture at 45°C on cellulose-containing media, supplemented with exogenous cellulase activity. At various stages during fermentation, both substrate and enzyme were added in batch mode and fermentation was continued for 220 h. Ethanol production increased to 20 g/l at 200 h, representing 45% of the maximum theoretical yield. In subsequent experiments, the organism was immobilized in calcium alginate beads and these were used in a similar, batch-fed system at 45°C. Again, fermentation was continued for 220 h and ethanol production increased to its maximum, of 28 g/l, within 100 h and this represented in excess of 60% of the maximum theoretical yield.
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    Structural and functional characterization of a stable, 4-chlorosalicylic-acid-degrading, bacterial community in a chemostat (1995)
    Springer
    World journal of microbiology and biotechnology 11 (1995), S. 643-645 
    Details
    ISSN: 1573-0972
    Keywords: Chemostat ; 4-chlorosalicylic acid ; continuous culture ; degradation ; microbial consortium
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract A mixed, stable microbial community, obtained by continuous enrichment of a sediment core using 4-chlorosalicylic acid as sole source of carbon and energy, contained 10 different bacterial species, including Klebsiella pneumonia, Pseudomonas fluorescens, P. mendocina and P. cichorii. The members of the community were grown separately on various chlorinated compounds which were readily degraded.
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    Cell-recycle batch fermentation using immobilized cells of flocculent Saccharomyces cerevisiae wine strains (1996)
    Springer
    World journal of microbiology and biotechnology 12 (1996), S. 25-27 
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    ISSN: 1573-0972
    Keywords: Batch fermentation ; immobilization ; Saccharomyces cerevisiae ; secondary products ; wine yeast ; wine making
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Five, highly flocculeng strains of Saccharomyces cerevisiae, isolated from wine, were immobilized in calcium alginate beads to optimize primary must fermentation. Three cell-recycle batch fermentations (CRBF) of grape musts were performed with the biocatalyst and the results compared with those obtained with free cells. During the CRBF process, the entrapped strains showed some variability in the formation of secondary products of fermentation, particularly acetic acid and acetaldehyde. Recycling beads of immobilized flocculent cells is a good approach in the development and application of the CRBF system in the wine industry.
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    URL: http://dx.doi.org/10.1007/BF00327794
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    Short Communication: Immobilization of urease from pigeonpea (Cajanus cajan L.) on flannel cloth using polyethyleneimine (1998)
    Springer
    World journal of microbiology and biotechnology 14 (1998), S. 927-929 
    Details
    ISSN: 1573-0972
    Keywords: Urease ; pigeonpea ; Cajanus cajan ; immobilization ; urea analysis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Urease of pigeonpea has been immobilized on polyethyleneimine-activated cotton cloth followed by cross-linking with dimethyl suberimidate. Optimum immobilization (56%) was obtained at a protein loading of 1.2mg/5×5cm2 cloth piece. The immobilized enzyme stored in 0.1M Tris/acetate buffer, pH6.5, at 4°C had a t1/2 of 70 days. There was practically no leaching of the enzyme from the immobilization matrix in 15 days. The immobilized enzyme was used 7 times at an interval of 24h between each use with 75% residual activity at the end of the period. Blood urea analysis was carried out with immobilized urease for some clinical samples.
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    URL: http://dx.doi.org/10.1023/A:1008897600256
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    Continuous cultivation of the yeast Candida utilis at different dilution rates on pineapple cannery effluent (1999)
    Springer
    World journal of microbiology and biotechnology 15 (1999), S. 115-117 
    Details
    ISSN: 1573-0972
    Keywords: Candida utilis ; pineapple cannery effluent ; continuous culture ; steady-state
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Candida utilis was grown on a pineapple cannery effluent in a chemostat at dilution rates ranging between 0.05 and 0.65 h−1 to establish optimal conditions for biomass production and chemical oxygen demand (COD) reduction. Sucrose, fructose and glucose were the main sugars in the effluent. Maximum value for cell yield coefficient and productivity were (0.686, gx/gs) and (2.96, gx/l/h) at a dilution rate of 0.425 and 0.475 h−1, respectively, while maximum COD reduction (98%) was attained at a dilution rate of 0.1 h−1. The maintenance coefficient attained a value of (0.093, gs/gx/h). An increase in dilution rate produced a higher protein content of the biomass.
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    Bioreduction of a carbon–nitrogen double bond using immobilized baker's yeast’a first report (1997)
    Springer
    World journal of microbiology and biotechnology 14 (1997), S. 247-250 
    Details
    ISSN: 1573-0972
    Keywords: Baker's yeast ; 18-crown-6 ; imines ; immobilization ; oximes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Immobilized baker's yeast entrapped in calcium alginate beads efficiently reduces N-benzylidinemethylamine to N-methylbenzylamine in hexane at 37°C and tetrahydrofuran (THF) at 30°C in the presence of 18-crown-6, while in the presence of water as cosolvent and glucose as an additive N-benzylidinemethylamine undergoes decomposition. Benzaldoxime in a hexane–water (1:9) solvent system containing glucose as an additive is reduced to N-benzylhydroxylamine. On using an ethanol–water (1:1) solvent system, benzaldoxime is converted to benzyl alcohol and in hexane, benzene, THF, hexane–water (1:1) or acetonitrile–water (1:1) solvent systems, or using dried baker's yeast in different solvent systems, transformation of benzaldoxime does not occur.
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    Removal of gaseous n-valeric acid in the air by Rhodococcus sp. B261 immobilized onto ceramic beads (1998)
    Springer
    World journal of microbiology and biotechnology 14 (1998), S. 343-348 
    Details
    ISSN: 1573-0972
    Keywords: Biofilter ; immobilization ; malodour ; volatile fatty acids
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract n-Valeric acid, one of the main malodorous pollutants from livestock houses was eliminated with a biofilter prepared with Rhodococcus sp. B261 immobilized onto ceramic beads. The strain was isolated from composted pig faeces and grown in an artificial medium containing volatile fatty acids as a carbon source. The cells were immobilized onto ceramic beads in vacuo. The beads were aseptically incubated at 37 °C, pH 8.0, for 24h for activation of the cells. The beads with immobilized cells (3.36×109 c.f.u./g ceramic beads) and moisture content of 35% (w/w) were packed into a glass column equipped with a water jacket to keep the temperature constant. One hundred-seventy ppm of gaseous n-valeric acid were removed for 11 days at 30h -1 (space velocity) and 37 °C.
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    Maintenance of steroid 11-hydroxylation activity in immobilized Cunninghamella elegans protoplasts (1997)
    Springer
    World journal of microbiology and biotechnology 13 (1997), S. 469-473 
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    ISSN: 1573-0972
    Keywords: 2-Deoxy-d-glucose ; hydroxylation ; immobilization ; polyoxin ; protoplasts ; steroids
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
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    URL: http://dx.doi.org/10.1023/A:1018540620234
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    Simultaneous filtration and immobilization of cells from a flowing suspension using a bioreactor containing polyethylenimine coated cotton threads: Application in the continuous inversion of concentrated sucrose syrups (1999)
    Springer
    World journal of microbiology and biotechnology 15 (1999), S. 17-21 
    Details
    ISSN: 1573-0972
    Keywords: Adhesion ; cotton threads ; immobilization ; invert sugar ; microbial filter ; polyethylenimine
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Polyethylenimine(PEI)-coated cotton threads were shown to have potential for reducing microbial load from a flowing suspension. Turbid cell suspensions perfused through the PEI column appeared as totally clear in the effluent. The adhesion efficiency of the matrix was found to depend on the concentration of PEI used to treat the threads. Threads coated with 2.5% PEI were found to show optimal retention of cells. A considerable amount of binding was seen over a broad range of ionic concentration (0–0.3 M) and pH (3.6–10.3). Under similar conditions control threads did not show any filtration capacity. Saccharomyces cerevisiae, Saccharomyces fragilis, Escherichia coli and an Acetobacter species could be effectively filtered using PEI-coated threads. This technique can find potential for the simultaneous filtration and immobilization of cells in a bioreactor to be used in continuous bioprocessing as exemplified for the inversion of sucrose syrups using baker's yeast. The bioreactor could continuously hydrolyse 60% (w/v) sucrose syrups with a productivity of 2.25 kg/day for over a month without loss in efficiency.
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    Reassessment of the bioenergetic yield of Arthrospira platensis using continuous culture (1999)
    Springer
    World journal of microbiology and biotechnology 15 (1999), S. 235-238 
    Details
    ISSN: 1573-0972
    Keywords: Arthrospira platensis ; bioenergetic yield ; continuous culture ; irradiance ; specific growth rate ; mixotrophy
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Reassessement of bioenergetic growth yield of Arthrospira platensis was performed by using continuous culture under both autotrophic and mixotrophic conditions. Continuous culture was carried out at dilution rates of 0.017, 0.023 and 0.030 h−1. Under these dilution rates bioenergetic yields ranged between 4.45–6.03 × 10−3 g biomass kJ−1 and between 5.42–7.46 × 10−3 g biomass kJ−1, under autotrophic and mixotrophic conditions respectively. A maximum bioenergetic yield of 8.1 × 10−3 g biomass kJ−1 using an autotrophic culture can be calculated. Pigment accumulation (chlorophyll a and carotenoids) may be related to light irradiance, reaching a maximum pigment concentration under light saturation irradiance. Phycocyanin concentration increased during light limitation.
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    Increased H2 production by immobilized microorganisms (1995)
    Springer
    World journal of microbiology and biotechnology 11 (1995), S. 156-159 
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    ISSN: 1573-0972
    Keywords: Bacillus ; biogas-H ; hydrogen ; immobilization ; mixed culture ; recycling
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Viable cells of H2-producers (Bacillus licheniformis and a mixed microbial culture) were immobilized on brick dust and in calcium alginate beads. In batch culture, cells of the mixed culture in the free state yielded 8.2 l H2/mol glucose utilized, whereasB. licheniformis evolved 13.1 l H2. Immobilized cells, however, gave 4-fold more H2 than the free bacteria. Highest yields were from the cells immobilized on brick dust. High H2-production rates continued over two rounds of re-use of the immobilized cells.
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    Determination of the respiration quotient in mammalian cell culture in bicarbonate buffered media (1995)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 45 (1995), S. 524-535 
    Details
    ISSN: 0006-3592
    Keywords: respiration quotient ; carbon dioxide evolution rate ; continuous culture ; cell metabolism ; bicarbonate buffer ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The determination of the respiration quotient (RQ = CER/OUR) has not been used so far as a tool for understanding animal cell metabolism. This is due to problems in measuring the carbon dioxide evolution rate (CER) rather than the oxygen uptake rate (OUR). The determination of the CER is complicated by the use of bicarbonate in the medium. Using liquid and gas balances we have derived an equation for continuous culture to quantify the amount of CO2 that comes from the bicarbonate in the feed. Under cell-free conditions, values predicted by this equation agree within 4% with the experimental results. In continuous culture using hybridoma cells, the CO2 from the feed, as determined by an IR-gas analyzer, was found to represent a significant amount of the total measured CO2 in the off-gas (50% in a suboptimal, and 30% in high-growth medium). Furthermore, the problem of CO2 loss from the medium during medium preparation and storage was solved using both a theoretical and an experimental approach. RQ values in continuous culture were evaluated for two different growth media. Small but significant differences in RQ were measured, which were matched by differences in specific antibody rates and other metabolic quotients. In a medium with Primatone RL, an enzymatic hydrolysate of animal cell tissue that causes a more than twofold increase in cell density, the RQ was found to be 1.05, whereas in medium without Primatone RL (but containing amino acids equivalent in composition and concentration to Primatone RL) the RQ was found to be 0.97. We suggest the RQ to be a useful parameter for estimating the physiological state of cells. Its determination could be a suitable tool for both the on-line control of animal cell cultivations and the understanding of cell metabolism. © 1995 John Wiley & Sons, Inc.
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    Synthesis of protein-containing polymers in organic solvents (1995)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 45 (1995), S. 10-17 
    Details
    ISSN: 0006-3592
    Keywords: proteins ; enzymes ; immobilization ; biopolymers ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Subtilisin has been modified with polyethylene glycol (PEG) monomethacrylate (MW 8000) by reductive alkylation, and incorporated into polymethyl methacrylate durring free-radical initiated polymerization. The activity and stability of the PEG-modified enzymes have been determined in aqueous buffer and organic solvents. The Km and Vmax values for unmodified, singly and doubly modified subtilisin were compared in these environments, and the half-lives of both modified enzymes were remarkably high (up to 2 months). The protein-containing polymer was analyzed for activity and polymer properties, and our results indicate that active subtilisin can be incorporated into polymethyl methacrylate during polymerization in organic solvents while retaining its activity and stability. © 1995 John Wiley & Sons, Inc.
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    Removal of phenols and aromatic amines from wastewater by a combination treatment with tyrosinase and a coagulant (1995)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 45 (1995), S. 304-309 
    Details
    ISSN: 0006-3592
    Keywords: phenol ; substituted phenol ; tyrosinase ; immobilization ; chitosan ; coagulant ; immobilized enzyme ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Removal of phenols and aromatic amines from industrial wastewater by tyrosinase was investigated. A color change from colorless to darkbrown was observed, but no precipitate was formed. Colored products were found to be easily removed by a combination treatment with tyrosinase and a cationic polymer coagulant containing amino group, such as hexamethylenediamine-epichlorohidrin polycondensate, polyethleneimine, or chitosan. The first two coagulants, synthetic polymers, were more effective than chitosan, a polymer produced in crustacean shells. Phenols and aromatic amines are not precipitated by any kind of coagulants, but their enzymatic reaction products are easily precipitated by a cationic polymer coagulant. These results indicate that the combination of tyrosinase and a cationic polymer coagulant is effective in removing carcinogenic phenols and aromatic amines from an aqueous solution. Immobilization of tyrosinase on magnetite gave a good retention of activity (80%) and storage stability i.e., only 5% loss after 15 days of storage at ambient temperature. In the treatment of immobilized tyrosinase, colored enzymatic reaction products were removed by less coagulant compared with soluble tyrosinase. © 1995 John Wiley & Sons, Inc.
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    Working at controlled water activity in a continuous process: The gas/solid system as a solution (1995)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 45 (1995), S. 387-397 
    Details
    ISSN: 0006-3592
    Keywords: transesterification ; water activity ; lipolytic enzymes ; gas ; bioreactor ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Fusarium solani cutinase and Candida cylindracea lipase were used to catalyze a transesterification reaction in a continuous gas/solid bioreactor. In this system, a solid phase composed of a packed enzymatic preparation was continuously percolated with carrier gas which fed substrate and removed reaction products simultaneously. Different conditions of immobilization were used and compared to the results obtained with a nonsupported enzyme. The enzymatic activity was found to be highly dependent of a key parameter: water activity (aw). Biocatalyst stability was greatly influenced by water activity and the choice of immobilization technique for the enzymatic material. For free and adsorbed enzymes, water requirements exhibited optima which corresponded to the complete hydration coverage of the protein. These optima presented a good correlation with the isotherm sorption curves obtained for the different preparations. In this work are reported the results concerning the possibility of using a continuous system able to operate at controlled water activity in a heterogeneous medium. Lipolytic enzyme in such a system appears to be a new process for the biotransformation of volatile esters. © 1995 John Wiley & Sons, Inc.
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    Sustained and constitutive high levels of protein production in continuous cultures of bacillus subtilis (1995)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 47 (1995), S. 520-524 
    Details
    ISSN: 0006-3592
    Keywords: bacillus subtilis ; plasmid ; continuous culture ; CAT ; recombinant cultures ; acid formation ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The feasibility of continuous production of proteins in chemostat cultures of Bacillus subtilis was investigated. An expression system consisting of the bacterium B. subtilis BR151 carrying plasmid p602/19 was used. The plasmid contains the cat (chioramphenicol acetyltrans-ferase) gene downstream of a strong vegetative T5 promoter. It was found that, at a dilution rate of 0.2 h-1 production of relatively high levels of CAT protein (about 4% ofcellular protein) can be sustained. But, experiments at a higher dilution rate of 0.4 h-1 were unproductive because of high acidformation and washout. Combination of low cell yield, which results from excessive acid formation, and low dilution rate led to a low volumetric CAT productivity. Our recent work with the nonrecombinant cells, has demonstrated that uptake of small amounts of citrate significantly reduces or entirelyeliminates the acid formation. This superior performance in the presence ofcitrate was hypothesized, based on strong experimental evidence, to be the result of a reduction in glycolysis flux through a sequence of events leading to a reduction in pyruvate kinase and phosphof- ructokinase activities, the regulatory enzymes of glycol-ysis. In this study, it is demonstrated that cofeeding of glucose and citrate substantially reduces theorganic acid formation and significantly increases the recombinant culture productivity. The combination of high specific CAT activity and cell density resulted in a total of six- to tenfold higher culture productivitywhen citrate and glucose were cometabolized than when glucose was the only carbon source. © 1995 John Wiley & Sons Inc.
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    Implantable hollow fiber bioreactor as a potential treatment for hypercholesterolemia: Characterization of the catalytic unit (1995)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 48 (1995), S. 36-41 
    Details
    ISSN: 0006-3592
    Keywords: hollow fiber ; bioreactor ; immobilized enzymes ; porosity ; phospholipase A2 ; low densitylipoprotein ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Previous studies have shown that the modification of low density lipoprotein (LDL) by the enzyme phospholipase A2(PLA2)results in a reduction of cholesterol levels in the plasma of hypercholesterolemic rabbits, due to accelerated clearance of the modified LDL. In the current study, we established techniques and optimized the ratio of enzyme to support for the immobilization of PLA2 on a polymeric support. Hollow fiber bioreactors made from polytetrafluoroethylene (PTFE) polymers were used to encapsulate immobilized PLA2. This design was adopted to eliminate hemolysis of red blood cells by the enzyme. Characterization of the resulting immobilized enzyme in terms of its activity, Michaelis-Menten kinetic constants, and the variation of its activity with incubation time is presented. The enzyme activity was not significantly altered upon incubation at 37°C in lipoprotein-deficient serum (LPDS), over the course of 2 months. The Michaelis-Menten kinetics constants are KM = 8.9 mM, Vmax = 6434.2 for the free enzyme and KappM = 16.7 mM, Vappmax = 619.7 for the immobilized enzyme. These data suggest that a system based on immobilized PLA2 in conjunction with hollow fiber bioreactors (HFBs) may be a good candidate for lowering LDL levels in plasma. © 1995 John Wiley & Sons, Inc.
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    Photochemistry of a photosynthetic reaction center immobilized in lipidic cubic phases (1995)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 46 (1995), S. 93-98 
    Details
    ISSN: 0006-3592
    Keywords: photosynthetic reaction center ; liquid crystals ; cubic phases ; immobilization ; Chloroflexus aurantiacus ; photochemistry ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Photosynthetic reaction centers, isolated and purified from the facultative phototrophic bacterium Chloroflexus aurantiacus, were immobilized in optically transparent lipidic cubic phases composed of 42% (w/w) 1-palmitoyl-2-hydroxy-sn-glycero-3-phosphocholine and 58% (w/w) water. The immobilized photosynthetic protein retains its native properties, as indicated by visible and circular dichroic spectra. The ground state visible spectrum of the immobilized reaction centers is very similar to the corresponding spectrum in aqueous solution, indicating that the protein pigments are not extracted into the lipidic regions of the cubic phase. The secondary structure of the protein is maintained in the immobilized state, as determined by far-UV circular dichroism spectroscopy in the 200- to 250-nm range. Moreover, immobilized reaction centers retain their photochemical activity: a reversible photo-oxidation of the primary electron donor (P) is seen upon continuous illumination. Furthermore, the entrappment of reaction centers does not affect the kinetics of charge recombination between the photo-oxidized primary donor (P+) and the photoreduced primary quinone acceptor, generated by a short flash of light. Reaction centers devoided of the secondary quinone acceptor can be easily reconstituted in cubic phases by means of their coimmobilization with 1,4-naphtoquinone. Indeed, the kinetics for charge recombination in reconstituted reaction centers is dramatically slower than the corresponding kinetics in the unreconstituted protein. Interestingly, immobilized reaction centers are significantly stabilized as compared with reaction centers in aqueous solution: the integrity of the protein in the cubic phase is maintained for at least 5 months, whereas in water solution 50% of the activity is lost within 2 months. © 1995 John Wiley & Sons, Inc.
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    Serum-free cell culture on insulin-immobilized porous collagen beads (1995)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 45 (1995), S. 144-148 
    Details
    ISSN: 0006-3592
    Keywords: serum-free cell culture ; cell adhesion ; cell growth ; fibroblast cell ; biosignal ; immobilization ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Insulin or albumin was immobilized on collagen beads using water-soluble carbodiimide. Adhesion of STO mouse fibroblast cells onto the beads decreased with increasing the amount of immobilized proteins. Growth of the cells was remarkably accelerated on the insulinimmobilized collagen beads, which can be used for serum-free cell culture. The growth acceleration became larger with increasing the amount of immobilized insulin, while it became smaller with increasing the amount of immobilized albumin. In addition, the immobilized insulin more strongly accelerated the cell growth than free insulin plus collagen beads. © 1995 John Wiley & Sons, Inc.
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    Protein-free human-human hybridoma cultures in an intercalated-spiral alternate-dead-ended hollow fiber bioreactor (1995)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 47 (1995), S. 384-400 
    Details
    ISSN: 0006-3592
    Keywords: hollow fiber ; bioreactor ; hybridoma ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Batch cell cultures of a human-human hybridoma line in a convective flow dominant intercalated-spiral altetnate-dead-ended hollow fiber are compared with those using conventional axial-flow hollow fiber bioreactors and a stirred-tank bioreactor. Relatively short-term fed-batch and perfusion cell cultures were also employed for the intercalated-spiral bioreactor. When operating conditions of a batch intercalated-spiral bioreactor were properly chosen, the cell growth and substrate consumption paralleled that of a batch stirred-tank culture. The results verified the premise of the intercalated-spiral hollow fiber bioreactor that nutrient transport limitations can be eliminated when the convective flux through the extracapillary space is sufficiently high.© John Wiley & Sons, Inc.
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    Studies on the lipozyme-catalyzed synthesis of butyl laurate (1995)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 46 (1995), S. 1-12 
    Details
    ISSN: 0006-3592
    Keywords: Lipozyme ; esterification ; immobilization ; butanol ; lauric acid ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The effects of temperature, speed of agitation, enzyme concentration, etc., on butyl laurate synthessis using Mucor miehei lipase (Lipozyme™) have been studied. Although the soluble enzyme was quite thermcstable in aqeous solution, it deactivated rapidly at and above 40°C in the presence of butanol. This enzyme immobilized on an anion-exchange resin (Lipozyme™) showed enhanced stability (as compared to the soluble form) to denaturation by butanol under the same conditions. The denaturation of M. miehei lipase was found to be a function of the butanol concentration in the aqueous phase, and rapid denaturation takes place at the concentration corresponding to its saturation at that temperature. © 1995 John Wiley & Sons, Inc.
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    Selective adsorption of proteins to their ligands covalently immobilized onto microfibers (1995)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 47 (1995), S. 557-566 
    Details
    ISSN: 0006-3592
    Keywords: polyester fiber ; immobilization ; protein A ; antigen ; antibody ; immunoadsorbent ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Following ozone oxidation of polyester microfibers of 3.5 μm average diameter and 0.83 m2/g specific area, the fiber surface was subjected to graft polymerization of acrylic acid and subsequently immobilized with serologically active proteins including Staphylococcus aureus protein A, a specific antigen, and a specific antibody. The immobilization reaction was mediated by a watersoluble carbodiimide, which allowed formation of a co-valent linkage between the ligand proteins and the grafted poly(acrylic acid)chains. The yields of the immobilized ligand proteins were of the order of 1 mg/g fiber. Their binding affinity and capacity to respective specific proteins were studied in vitro from a buffered solution and serum. It was found that the specific proteins were selectively adsorbed with dissociation constants as low as 1× 10-6 M, suggesting the adsorption to take place through highly specific protein-protein interaction. An addition of serum albumin did not significantly affect the specific binding, regardless of the ligand proteins. The binding capacity ranged from 1 × 10-13 to 1× 10-11 mol/cm2 primarily depending on the surface density of the immobilized ligands and the number of their binding sites per molecule. © 1995 John Wiley & Sons Inc.
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    Importance of spore mutants for fed-batch and continuous fermentation of Bacillus subtilis (1995)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 47 (1995), S. 696-702 
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    ISSN: 0006-3592
    Keywords: Bacillis subtilis ; spore mutant ; fed-batch ; continuous culture ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: To alleviate plasmid instability and to prolong the production phase of subtilisin, integrable plasmid and spore mutants are used. Compared with batch-type shake flask cultures, spore mutants' ability to produce subtilisin can be well pronounced in fed-batch and continuous cultures. Hence, the two culture methods make it possible to identify the peculiar characteristics of the spore mutants unobtainable in batch culture. Spore mutants can enhance subtilisin productivity and prolong subtilisin production time in fed-batch culture as well as enable us to use very low dilution rates (〈0.1 h-1) without losing productivity in continuous culture, thereby improving the conversion yield of the nitrogen source. At 0.05 h-1 the spollG mutant of Bacillus subtilis DB104 (Δnpr Δapr) (Emr) spollG (Bimr):: pMK101 (Cmr) showed a subtilisin yield about ten times higher than that from wild-type DB104 (Δnpr Δapr)::pMK101 (Cmr). © 1995 John Wiley & Sons, Inc.
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    A new continuous biofilm bioreactor for immobilized oil-degrading filamentous fungi (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 49 (1996), S. 20-25 
    Details
    ISSN: 0006-3592
    Keywords: filamentous fungi ; immobilization ; biofilm bioreactor ; oil emulsion ; degradation ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A new type of horizontal biofilm bioreactor for continuous bioconversion of emulsified oily substrate by immobilized growing biofilm of filamentous fungi was designed, constructed, and feasibility tested. The new reactor design provides “self”-immobilization of homogenized mycelium leading to even biofilm development. This was accomplished by using stainless steel screens of optimal mesh, mounted in parallel and stretching outward from a main rotating axis of a biological rotating contractor. Each screen was equipped with a pair of stainless steel blades mounted on supports allowing for continuous biofilm “shaving” beyond a predetermined thickness, thus retaining freshly growing active biofilm surface. The feasibility of the new bioreactor was demonstrated by decalactone production from emulsified castor oil by immobilized filamentous fungi (Tyromyces sambuceus). The combination of oriented metal screens and moving blades was found to be highly effective for a model system in maintaining stable substrate emulsion in the reactor in either batchwise or continuous processing, as well as maintaining biofilm thickness with continuous removal of excess growing hyphae. © 1996 John Wiley & Sons, Inc.
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    Microencapsulation of yeast cells and their use as a biocatalyst in organic solvents (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 49 (1996), S. 535-543 
    Details
    ISSN: 0006-3592
    Keywords: whole cell biotransformation ; biocatalyst ; baker's yeast ; immobilization ; microencapsulation ; organic solvents ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Stable, semipermeable polyamide microcapsules were prepared by interfacial polymerization from a mixture of 1,6-hexanediamine and poly(allylamine) crosslinked with di-acid chlorides and were used to encapsulate baker's yeast. The size and distribution of cells within the capsules were investigated by a combination of laser confocal, electron scanning, and transmission electron microscopy. The encapsulated cells were studied as a biocatalyst for the model reduction of 1-phenyl-1,2-propanedione to 2-hydroxy-1-phenyl-1-propanone in a number of organic solvents. The polymerization conditions were extensively investigated and were found to greatly influence the product yield. Microencapsulated yeast cells, prepared under optimized conditions, carried out the reduction more efficiently than free cells as well as those immobilized in alginate and κ-carrageenan beads. The developed methodology should be broadly applicable to other biotransformations of interest. © 1996 John Wiley & Sons, Inc.
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    Biological sulfate reduction using synthesis gas as energy and carbon source (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 50 (1996), S. 136-144 
    Details
    ISSN: 0006-3592
    Keywords: sulfate-reducing bacteria ; biofilm ; immobilization ; gas-lift reactor ; carbon monoxide ; synthesis gas ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Biological sulfate reduction was studied in laboratory-scale gas-lift reactors. Synthesis gas (gas mixtures of H2/CO/CO2) was used as energy and carbon source. The required biomass retention was obtained by aggregation and immobilization on pumice particles. Special attention was paid to the effect of CO addition on the sulfate conversion rate, aggregation, and aggregate composition.Addition of 5% CO negatively affected the overall sulfate conversion rate; i.e., it dropped from 12-14 to 6-8 g SO2-4/L day. However, a further increase of CO to 10 and 20% did not further deteriorate the process. With external biomass recycling the sulfate conversion rate could be improved to 10 g SO2-4/L day. Therefore biomass retention clearly could be regarded as the rate-limiting step. Furthermore, CO affected the aggregate shape and diameter. Scanning electron microscopy (SEM) photographs showed that rough aggregates pregrown on H2/CO2 changed into smooth aggregates upon addition of CO. Addition of CO also changed the aggregate Sauter mean diameter (d32) from 1.7 mm at 5% CO to 2.1 mm at 20% CO. After addition of CO, a layered biomass structure developed. Acetobacterium sp. were mainly located at the outside of the aggregates, whereas Desulfovibrio sp. were located inside the aggregates. © 1996 John Wiley & Sons, Inc.
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    Immobilization of Candida rugosa lipase to nylon fibers using its carbohydrate groups as the chemical link (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 51 (1996), S. 327-341 
    Details
    ISSN: 0006-3592
    Keywords: Candida rugosa ; immobilization ; olive oil hydrolysis ; phenylglycidate ; glutaraldehyde ; adipic dihydrazide ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The aim of this study was to evaluate the immobilization of lipase from Candida rugosa on a nylon support by methods used to attach biomolecules to solid supports through their carbohydrate moieties. The carbohydrate groups were converted to dialdehydes by treatment with sodium periodate. The length of exposure and the periodate amount were optimized to the point where almost total activity retention was obtained. Tests of the immobilized enzyme showed the expressed activity to be significantly higher than the activity obtained with the unimmobilized enzyme. The use of reverse micelles as a way of delivering water to the enzyme was tested and found to give significantly higher activities. The immobilized enzyme activity was also tested with other substrates, one of which was a chiral ester. The immobilized enzyme was found to have high stereoselective efficiency and activity toward racemic methyl methoxyphenyl glycidate, a chiral intermediate used in the manufacture of the drug diltiazem. © 1996 John Wiley & Sons, Inc.
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    Activation and regeneration of whole cell biocatalysts: Initial and periodic induction behavior in starved Escherichia coli after immobilization in thin synthetic films (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 51 (1996), S. 360-370 
    Details
    ISSN: 0006-3592
    Keywords: induction ; Escherichia coli ; biofilms ; immobilization ; protein synthesis in starved bacteria ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Activation and regeneration of whole cell biocatalytic activity via initial and subsequent induction of the lacZ gene was investigated in starved Escherichia coli using a novel synthetic biofilm. Stationary-phase bacteria were entrapped in 10-80 μm thick multi-layer films, where a copolymer of acrylic and vinyl acetate was the immobilization matrix. The E. coli were placed in a defined starvation medium containing essentially no nitrogen or carbon source and induced initially using lactose or isopropylthiogalactoside (IPTG). Subsequent inductions were performed with IPTG. Comparison studies with suspended bacteria showed that when IPTG was the initial inducing agent, induction kinetics are linear for both immobilized and suspended cells. After induction with lactose, however, a lag time is noted for suspended cells, but not for E. coli in the biofilm. Biocatalytic activity was successfully regenerated by re-inducing starved suspended cells 1-3 days after an initial induction with lactose. This regeneration was demonstrated in the synthesis of additional active β-galactosidase. However, immobilized cells could be re-induced for at least 17 days after the initial induction, and viability in the synthetic biofilms remained greater than 90%, demonstrating that periodic induction is a valuable method for extending the life of whole cell biocatalysts. © 1996 John Wiley & Sons, Inc.
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    Transient and stationary operating conditions on performance of lactic acid bacteria crossflow microfiltration (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 49 (1996), S. 78-86 
    Details
    ISSN: 0006-3592
    Keywords: crossflow microfiltration ; hydrodynamics ; fouling ; bioreactor ; Lactobacillus helveticus ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A filtration rig equipped with a tubular alumina membrane was used to study the performance of crossflow microfiltration of Lactobacillus helveticus. Experiments were performed at constant permeation flux. High cell concentrations and fast transient conditions to the stationary J adversely affected permeability. Membrane fouling was due to a fast irreversible layer formation and to a reversible cell cake. This microbial deposit characteristics were dependent on the ratio permeation flux/wall shear stress, J/τw. Fouling was faster and more severe when J/τw was greater than a critical value of 1.15 L-1 · h-1 · m-2 · Pa-1. The disordered structure of this cell cake seemed to lead to a macromolecule deposit between the cells which adversely affected the membrane permeability. © 1996 John Wiley & Sons, Inc.
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    Properties of free and immobilized lipase from Pseudomonas cepacia (1997)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 56 (1997), S. 181-189 
    Details
    ISSN: 0006-3592
    Keywords: lipase ; immobilization ; polypropylene support ; Pseudomonas cepacia ; kinetic parameters ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The purified lipase from Pseudomonas cepacia (PS, Amano) was immobilized on a commercially available microporous polypropylene support. The enzyme was rapidly and completely adsorbed on the support. Special attention was devoted to the demonstration of the lack of diffusional limitations, either internal or external, when a soluble substrate (p-nitrophenylacetate, pNPA) was used. The activity yield was high (100%) with pNPA and very low (0.4%) with p-nitrophenylpalmitate (pNPP). These values clearly showed that the immobilized enzyme was fully active as soon as activity was assayed on a soluble substrate rather than an insoluble one. With the latter one, the low activity was due mainly to a slow rate of substrate diffusion inside the porous support. The same diffusional phenomenon could explain the complete change of fatty acid specificity of the immobilized lipase. After immobilization, the lipase was mainly specific for short chain fatty acid esters, whereas the free enzyme was mainly specific for long chain esters. The activity-versus-temperature profiles were not greatly affected by immobilization with maximal reaction rates in the range 45° to 50°C for both enzyme preparations. However, immobilization increased enzyme stability mainly by decreasing the sensitivity to temperature of the inactivation reaction. Half-lives at 80°C were 11 and 4 min for the immobilized and free enzymes, respectively. © 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 56: 181-189, 1997.
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    Cell growth and differentiation on feeder layers is predicted to be influenced by bioreactor geometry (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 50 (1996), S. 479-492 
    Details
    ISSN: 0006-3592
    Keywords: stem cell ; bioreactor ; stromal layer ; Graetz number ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Tissue function is comprised of a complex interplay between biological and physicochemical rate processes. The design of bioreactors for tissue engineering must account for these processes simultaneously in order to obtain a bioreactor that provides a uniform environment for tissue growth and development. In the present study we consider the effects of fluid flow and mass transfer on the growth of a tissue in a parallel-plate bioreactor configuration. The parenchymal cells grow on a preformed stromal (feeder) layer that secretes a growth factor that stimulates parenchymal stem cell replication and differentiation. The biological dynamics are described by a unilineage model that describes the replication and differentiation of the tissue stem cell. The physicochemical rates are described by the Navier-Stokes and convective-diffusion equations. The model equations are solved by a finite element method. Two dimensionless groups govern the behavior of the solution. One is the Graetz number (Gz) that describes the relative rates of convection and diffusion, and the other a new dimensionless ratio (designated by P) that describes the interplay of the growth factor production, diffusion, and stimulation. Four geometries (slab, gondola, diamond, and radial shapes) for the parallel-plate bioreactor are analyzed. The uniformity of cell growth is measured by a two-dimensional coefficient of variance. The concentration distribution of the stroma-derived growth factor was computed first based on fluid flow and bioreactor geometry. Then the concomitant cell density distribution was obtained by integrating the calculated growth factor concentration with the parenchymal cell growth and unilineage differentiation process. The spatiotemporal cell growth patterns in four different bioreactor configurations were investigated under a variety of combinations of Gz (10-1, 100, and 101) and P(10-2, 10-1, 100, 101, and 102). The results indicate high cell density and uniformity can be achieved for parameter values of P = 0.01, …, 0.1 and Gz = 0.1, …, 1.0. Among the four geometries investigated the radial-flow-type bioreactor provides the most uniform environment in which parenchymal cells can grow and differentiate ex vivo due to the absence of walls that are parallel to the flow paths creating slow flowing regions. © 1996 John Wiley & Sons, Inc.
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    Hollow fiber bioartificial liver utilizing collagen-entrapped porcine hepatocyte spheroids (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 52 (1996), S. 34-44 
    Details
    ISSN: 0006-3592
    Keywords: hepatocyte spheroid ; porcine hepatocyte ; hollow fiber ; bioartificial liver ; collagen ; bioreactor ; ureagenesis ; albumin synthesis ; glucuronidation ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A xenogeneic hollow fiber bioreactor utilizing collagen-entrapped dispersed hepatocytes has been developed as an extracorporeal bioartificial liver (BAL) for potential treatment of acute human fulminant hepatitis. Prolonged viability, enhanced liver-specific functions, and differentiated state have been observed in primary porcine hepatocytes cultivated as spheroids compared to dispersed hepatocytes plated on a monolayer. Entrapment of spheroids into the BAL can potentially improve performance over the existing device. Therefore, studies were conducted to evaluate the feasibility of utilizing spheroids as the functionally active component of our hybrid device. Confocal microscopy indicated high viability of spheroids entrapped into cylindrical collagen gel. Entrapment of spheroids alone into collagen gel showed reduced ability to contract collagen gel. By mixing spheroids with dispersed cells, the extent of collagen gel contraction was increased. Hepatocyte spheroids collagen-entrapped into BAL devices were maintained for over 9 days. Assessment of albumin synthesis and ureagenesis within a spheroid-entrapment BAL indicated higher or at least as high activity on a per-cell basis compared to a dispersed hepatocyte-entrapment BAL device. Clearance of 4-methylumbelliferone to its glucuronide was detected throughout the culture period as a marker of phase II conjugation activity. A spheroid-entrapment bioartificial liver warrants further studies for potential human therapy. © 1996 John Wiley & Sons, Inc.
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    Biooxidation capacity of the extremely thermoacidophilic archaeon Metallosphaera sedula under bioenergetic challenge (1998)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 58 (1998), S. 617-624 
    Details
    ISSN: 0006-3592
    Keywords: thermoacidophile ; chemolithotroph ; heat shock ; chemical stress ; continuous culture ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The biooxidation capacity of an extremely thermoacidophilic archaeon Metallosphaera sedula (DSMZ 5348) was examined under bioenergetic challenges imparted by thermal or chemical stress in regard to its potential use in microbial bioleaching processes. Within the normal growth temperature range of M. sedula (70-79°C) at pH 2.0, upward temperature shifts resulted in bioleaching rates that followed an Arrhenius-like dependence. When the cells were subjected to supraoptimal temperatures through gradual thermal acclimation at 81°C (Han et al., 1997), cell densities were reduced but 3 to 5 times faster specific leaching rates (Fe3+ released from iron pyrite/cell/h) could be achieved by the stressed cells compared to cells at 79°C and 73°C, respectively. The respiration capacity of M. sedula growing at 74°C was challenged by poisoning the cells with uncouplers to generate chemical stress. When the protonophore 2,4-dinitrophenol (5-10 μM) was added to a growing culture of M. sedula on iron pyrite, there was little effect on specific leaching rates compared to a culture with no protonophore at 74°C; 25 μM levels proved to be toxic to M. sedula. However, a significant stimulation in specific rate was observed when the cells were subjected to 1 μM nigericin (+135%) and 2 μM (+63%); 5 μM levels of the ionophore completely arrested cell growth. The ionophore effect was further investigated in continuous culture growing on ferrous sulfate at 74°C. When 1 μM nigericin was added as a pulse to a continuous culture, a 30% increase in specific iron oxidation rate was observed for short intervals, indicating a potential positive impact on leaching when periodic chemical stress is applied. This study suggests that biooxidation rates can be increased by strategic exposure of extreme thermoacidophiles to chemical or thermal stress, and this approach should be considered for improving process performance. © 1998 John Wiley & Sons, Inc. Biotechnol Bioeng 58: 617-624, 1998.
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    Performance of a sulfide-oxidizing expanded-bed reactor supplied with dissolved oxygen (1997)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 53 (1997), S. 32-40 
    Details
    ISSN: 0006-3592
    Keywords: expanded-bed reactor ; sulfur ; Thiobacilli ; immobilization ; biofilm ; sludge ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The performance of a new sulfide-oxidizing, expanded-bed bioreactor is described. To stimulate the formation of well-settleable sulfur sludge, which comprises active sulfide-oxidizing bacterial biomass and elemental sulfur, the aeration of the liquid phase and the oxidation of sulfide to elemental sulfur are spatially separated. The liquid phase is aerated in a vessel and subsequently recirculated to the sulfide-oxidizing bioreactor. In this manner, turbulencies due to aeration of the liquid phase in the bioreactor are avoided. It appeared that, under autotrophic conditions, almost all biomass present in the reactor will be immobilized within the sulfur sludge which consists mainly of elemental sulfur (92%) and biomass (2.5%). The particles formed have a diameter of up to 3 mm and can easily be grinded down. Within time, the sulfur sludge obtained excellent settling properties; e.g., after 50 days of operation, 90% of the sludge settles down at a velocity above 25 m h-1 while 10% of the sludge had a sedimentation velocity higher than 108 m h-1. Because the biomass is retained in the reactor, higher sulfide loading rates may be applied than to a conventional “free-cell” suspension. The maximum sulfide-loading rate reached was 14 g HS- L-1 d-1, whereas for a free-cell suspension a maximum loading rate of 6 g HS- L-1 d-1 was found. At higher loading rates, the upward velocities of the aerated suspension became too high so that sulfur sludge accumulated in the settling zone on top of the reactor. When the influent was supplemented with volatile fatty acids, heterotrophic sulfur and sulfate reducing bacteria, and possibly also (facultatively) heterotrophic Thiobacilli, accumulated within the sludge. This led to a serious deterioration of the system; i.e., the sulfur formed was increasingly reduced to sulfide, and also the formation rate of sulfur sludge declined. © 1997 John Wiley & Sons, Inc.
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    Study of stability of recombinant plasmids during the continuous culture of Bacillus stearothermophilus NUB3621 in nonselective medium (1997)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 53 (1997), S. 507-514 
    Details
    ISSN: 0006-3592
    Keywords: Bacillus stearothermophilus ; continuous culture ; plasmid stability ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The optimal culture conditions for Bacillus stearothermophilus NUB3621 (BGSC 9A5) in chemostat were studied. The results obtained showed that the optimal culture conditions in terms of biomass concentration and maximum growth rate were 65°C, pH 6.8 to 7.2. Dissolved oxygen became growth limiting at pO2 levels below 10%. Furthermore, this strain was transformed with three new hybrid vectors (pPAM2, pPCH2, or pPLY2) constructed by cloning in pRP9, a plasmid based on the thermophilic replicon, pBC1, and three heterologous genes: the α-amylase gene from Bacillus licheniformis, the cholesterol oxidase gene from Streptomyces sp., and the lipase gene from Pseudomonas fluorescens. The influence of several fermentative conditions on segregational and structural stability of the recombinant B. stearothermophilus NUB3621 transformants was studied.The parameters of plasmid loss, that is, rate of plasmid loss (R) and specific growth rate difference (δμ), were calculated. B. stearothermophilus NUB3621 carrying pRP9 showed great segregational stability in all the assayed conditions, exceeding more than 300 generations without significant plasmid loss, whereas NUB3621 carrying pPAM2, pPCH2, or pPLY2 exhibited relatively low plasmid stability. The segregational instability of the recombinant constructs increased by increasing the fermentation temperature, decreased by increasing the dilution rate, and was not affected by the level of dissolved oxygen. On the other hand, plasmid maintenance decreased in minimal medium if compared with the results obtained in complex medium. Restriction analyses carried out on cultures of NUB3621 carrying pRP9, pPAM2, pPCH2, or pPLY2, grown for 200 generations on nonselective media, revealed that all the clones tested contained the parental plasmids. These results indicate that the heterologous inserts did not affect the structural stability of the recombinant plasmids. © 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 53: 507-514, 1997.
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    Biodegradation of paint stripper solvents in a modified gas lift loop bioreactor (1997)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 55 (1997), S. 163-169 
    Details
    ISSN: 0006-3592
    Keywords: bioreactor ; paint stripper solvents ; biodegradation ; model ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Paint stripping wastes generated during the decontamination and decommissioning of former nuclear facilities contain paint stripping organics (dichloromethane, 2-propanol, and methanol) and bulk materials containing paint pigments. It is desirable to degrade the organic residues as part of an integrated chemical-biological treatment system. We have developed a modified gas lift loop bioreactor employing a defined consortium of Rhodococcus rhodochrous strain OFS and Hyphomicrobium sp. DM-2 that degrades paint stripper organics. Mass transfer coefficients and kinetic constants for biodegradation in the system were determined. It was found that transfer of organic substrates from surrogate waste into the air and further into the liquid medium in the bioreactor were rapid processes, occurring within minutes. Monod kinetics was employed to model the biodegradation of paint stripping organics. Analysis of the bioreactor process was accomplished with BIOLAB, a mathematical code that simulates coupled mass transfer and biodegradation processes. This code was used to fit experimental data to Monod kinetics and to determine kinetic parameters. The BIOLAB code was also employed to compare activities in the bioreactor of individual microbial cultures to the activities of combined cultures in the bioreactor. This code is of benefit for further optimization and scale-up of the bioreactor for treatment of paint stripping and other volatile organic wastes in bulk materials. © 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 55: 163-169, 1997.
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    Continuous culture study of the expression of hepatitis B surface antigen and its self-assembly into virus-like particles in Saccharomyces cerevisiae (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 49 (1996), S. 578-586 
    Details
    ISSN: 0006-3592
    Keywords: Saccharomyces cerevisiae ; hepatitis B surface antigen (HBsAg) ; continuous culture ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: We have studied the growth rate dependence of hepatitis B surface antigen (HBsAg) p24s monomer and lipoprotein particle synthesis produced in Saccharomyces cerevisiae using galactose-limited continuous culture. The hepatitis B virus S gene, which encodes the p24s monomer, is transcribed under the control of the GAL 10p on a chimeric 2-μm plasmid harbored in a haploid yeast strain. Monomers autonomously form lipoprotein aggregates (particles) in vivo using only host-cell-derived components. Steady states were evaluated in a range from 0.015 h-1 to washout (0.143 h-1). Both p24s monomer and HBsAg particle levels, at steady state, varied in an inverse linear manner with growth rate. A consistent excess of total p24s monomer to HBsAg particle, estimated at five- to tenfold by mass, was found at all dilution rates. The average copy number of the 2-μm plasmid (carrying LEU2 selection) remained constant at 200 copies per cell from washout to 0.035 h-1. Surprisingly, the average copy number was undetectable at the lowest dilution rate tested (0.015 h-1), even though HBsAg expression was maximal. Total p24s monomer and HBsAg particle values ranged twofold over this dilution rate range. No differences in the trends for HBsAg expression and average copy number could be detected past the critical dilution rate where aerobic fermentation of galactose and ethanol overflow were observed. HBsAg expression in continuous culture was stable for at least 40 generations at 0.100 h-1. © 1996 John Wiley & Sons, Inc.
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    Enhancement of monoclonal antibody production by immobilized hybridoma cell culture with hyperosmolar medium (1995)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 48 (1995), S. 699-705 
    Details
    ISSN: 0006-3592
    Keywords: hybridoma ; hyperosmotic stress ; immobilization ; antibody productivity ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: To determine the effect of hyperosmotic stress on the monoclonal antibody (MAb) production by calcium-alginate-immobilized S3H5/γ2bA2 hybridoma cells, the osmolalities of medium in the MAb production stage were varied through the addition of NaCI. The specific MAb productivity (qMAb) of immobilized cells exposed to abrupt hyperosmotic stress (398 mOsm/kg) was increased by 55% when compared with that of immobilized cells in the control culture (286 mOsm/kg). Furthermore, this enhancement of qMAb was not transient. Abrupt increase in osmolality, however, inhibited cell growth, resulting in no increase in volumetric MAb productivity (rMAb). On the other hand, gradual increase in osmolality allowed further cell growth while maintaining the enhanced qMAb immobilized cells. The qMAb immobilized cells at 395 mOsm/kg was 0.661 ± 0.019 μg/106 cells/h, which is almost identical to that of immobilized cells exposed to abrupt osmotic stress. Accordingly, the rMAb was increased by ca. 40% when compared with that in the control immobilized cell culture. This enhancement in iMAb of immobilized S3H5/γ2bA2 hybridoma cells by applying gradual osmotic stress suggests the potential of using hyperosmolar medium in other perfusion culture systems for improved MAb production. © 1995 John Wiley & Sons, Inc.
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    The viability of Scenedesmus bicellularis cells immobilized on alginate screens following nutrient starvation in air at 100% relative humidity (1995)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 46 (1995), S. 459-464 
    Details
    ISSN: 0006-3592
    Keywords: immobilization ; screens ; microalgae ; Scenedesmus bicellularis ; starvation in air ; relative humidity ; wastewater treatment ; nutrient uptake ; viability ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The viability of algal cells immobilized on screens and starved in a water-saturated air stream was studied at the laboratory scale. This new process for wastewater biotreatment has been developed using immobilized cells, which were starved in air, to obtain a high rate of nutrient removal. A unicellular green microalgae, Scenedesmus bicellularis, was isolated from secondary decantation tanks at an urban wastewater treatment plant and grown in a synthetic medium for 12 days. The cells were then concentrated by centrifugation and immobilized on alginate screens. The screens were then inserted in a photochamber saturated at 100% relative humidity and subjected to a photoperiod of 16 h in the light and 8 h in the dark, with an illumination of 150 μE m-2 s-1 provided by fluorescent lamps. After 48 h of nutrient starvation, the immobilized cells were used for the removal of ammonium and orthophosphate from a synthetic secondary wastewater effluent in a plexiglass reactor. During the sequential operation of starvation followed by incubation in the presence of nutrients, fast growth of viable cells in the gel matrix was obtained and there was no appreciable decay of chlorophyll a or cell activity. When these immobilized and starved cells were incubated in wastewater, ammonium (NH4+) and orthophosphate (PO43-) ions were quickly taken up from medium. After three successive 2-h exposures to wastewater, immobilized algal cells were freed by dissolving the Ca-alginate with phosphate as 0.2 M Na3PO4 and resuspended in fresh culture medium. Results indicate that free cells transferred to rich medium remained viable, but the growth rate revealed that the viable cells decreased their culturability. © 1995 John Wiley & Sons, Inc.
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    Growth kinetics of a bacteriophage in continuous culture (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 50 (1996), S. 217-221 
    Details
    ISSN: 0006-3592
    Keywords: Qβ phage ; molecular evolution ; phage display ; continuous culture ; cellstat ; wall growth ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Lytic coliphage Qβ was grown in continuously cultured host bacteria using a cascade of stirred flow reactors. The apparatus was constructed so that the steady stream of exponentially growing bacterial cells passing through the stirred flow reactors served to prevent coevolution brought about by host-parasite interactions. Wall growth was the primary cause for deviation from ideal continuous culture conditions and is largely dependent on the surface structure of the host bacteria. Using an Escherichia coli strain deficient in adhesive type I pili expression, the desynchronization of single burst events could easily be followed over the course of four infection latency periods. Computer simulations based on a two-stage model for the Qβ infection cycle were in perfect agreement with the experimental data. Applications of the optimized system to strategies of molecular evolution are discussed. © 1996 John Wiley & Sons, Inc.
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    Mixing power, external convection, and effectiveness in bioreactors (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 51 (1996), S. 131-140 
    Details
    ISSN: 0006-3592
    Keywords: mixing power ; convection ; fermentation ; bioreactor ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The phenomena of mixing and mass transfer of substrates to microorganisms greatly affect the biochemical reactions which take place in fermentation processes. The effect that agitation power has on the observable reaction kinetics involved in beer fermentation has been studied in different types of bioreactors, from laboratory to industrial scale. With this aim in mind, an effectiveness factor, η, is introduced which is defined as the relation between the existing rate of reaction, whichever bioreactor is considered, and the reaction rate in the well-mixed, and therefore presumably homogeneous, bioreactor with no diffusional limits. The limitation to homogeneously supplying nutrient material to the cells produces a decrease in this effectiveness factor, which has been correlated to the energy dissipation rate with a similar slope to that which appears in an existing correlation in the literature between this energy and the mass transfer coefficient. Additionally, a dimensionless reaction-convection number, NRC, which is a function of the power input per unit volume, is proposed, which has been appropriately employed in correlating the effectiveness factor for the types of processes in which convection may be the key resistance factor. © 1996 John Wiley & Sons, Inc.
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    Effect of particle loading on GM-CSF production by Saccharomyces cerevisiae in a three-phase fluidized bed bioreactor (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 51 (1996), S. 229-236 
    Details
    ISSN: 0006-3592
    Keywords: bioreactor ; fluidized bed ; murine granulocyte-macrophage colony stimulating factor ; Saccharomyces cerevisiae ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Continuous production of a recombinant murine granulocyte-macrophage colony-stimulating factor (MuGM-CSF) by immobilized yeast cells, Saccharomyces cerevisiae strain XV2181 (a/a, Trp1) containing plasmid pαADH2, in a fluidized bed bioreactor was studied at a 0.03 h-1 dilution rate and various particle loading rates ranging from 5% to 33% (v/v). Cells were immobilized on porous glass beads fluidized in an air-lift draft tube bioreactor. A selective medium containing glucose was used to start up the reactor. After reaching a stable cell concentration, the reactor feed was switched to a rich, nonselective medium containing ethanol as the carbon source for GM-CSF production. GM-CSF production increased initially and then dropped gradually to a stable level. During the same period, the fraction of plasmid-carrying cells declined continuously to a lower level, depending on the particle loading. The relatively stable GM-CSF production, despite the large decline in the fraction of plasmid-carrying cells, was attributed to cell immobilization. As the particle loading rate increased, the plasmid stability also increased. Also, as the particle loading increased from 5% to 33%, total cell density in the bioreactor increased from 16 to 36 g/L, and reactor volumetric productivity increased from 0.36 to 1.31 mg/L·h. However, the specific productivity of plasmid-carrying cells decreased from 0.55 to 0.07 mg/L·g cell. The decreased specific productivity at higher particle loading rates was attributed to reduced growth efficiency caused by nutrient limitations at higher cell densities. Both the reactor productivity and specific cell productivity increased by two- to threefold or higher when the dilution rate was increased from 0.03 to 0.07 h-1. © 1996 John Wiley & Sons, Inc.
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    Xylitol production by immobilized recombinant Saccharomyces cerevisiae in a continuous packed-bed bioreactor (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 51 (1996), S. 317-326 
    Details
    ISSN: 0006-3592
    Keywords: xylitol ; recombinant yeast ; immobilization ; continuous packed-bed reactor ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Continuous xylitol production with two different immobilized recombinant Saccharomyces cerevisiae strains (H475 and S641), expressing low and high xylose reductase (XR) activities, was investigated in a lab-scale packed-bed bioreactor. The effect of hydraulic residence time (HRT; 1.3-11.3 h), substrate/cosubstrate ratio (0.5 and 1), recycling ratio (0, 5, and 10), and aeration (anaerobic and oxygen limited conditions) were studied. The cells were immobilized by gel entrapment using Ca-alginate as support and the beads were treated with Al3+ to improve their mechanical strength. Xylose was converted to xylitol using glucose as cosubstrate for regeneration of NAD(P)H required in xylitol formation and for generation of maintenance energy. The stability of the recombinant strains after 15 days of continuous operation was evaluated by XR activity and plasmid retention analyses. Under anaerobic conditions the volumetric xylitol productivity increased with decreasing HRT with both strains. With a recycling ratio of 10, volumetric productivities as high as 3.44 and 5.80 g/L · h were obtained with the low XR strain at HRT 1.3 h and with the high XR strain at HRT 2.6 h, respectively. However, the highest overall xylitol yields on xylose and on cosubstrate were reached at higher HRTs. Lowering the xylose/cosubstrate ratio from 1 to 0.5 increased the overall yield of xylitol on xylose, but the productivity and the xylitol yield on cosubstrate decreased. Under oxygen limited conditions the effect of the recycling ratio on production parameters was masked by other factors, such as an accumulation of free cells in the bioreactor and severe genetic instability of the high XR strain. Under anaerobic conditions the instability was less severe, causing a decrease in XR activity from 0.15 to 0.10 and from 3.18 to 1.49 U/mg with the low and high XR strains, respectively. At the end of the fermentation, the fraction of plasmid bearing cells in the beads was close to 100% for the low XR strain; however, it was significantly lower for the high XR strain, particularly for cells from the interior of the beads. © 1996 John Wiley & Sons, Inc.
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    Covalent binding of a nerve agent hydrolyzing enzyme within polyurethane foams (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 51 (1996), S. 450-457 
    Details
    ISSN: 0006-3592
    Keywords: enzymes ; immobilization ; phosphotriesterase ; polyurethane foam ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A phosphotriesterase preparation, extracted from Escherichia coli DH5α cells, was immobilized within a polyurethane foam matrix during polymer synthesis. The enzyme-foam interaction was shown to be covalent and analysis of the hydrolysis of paraoxon in aqueous solution demonstrated that more than 50% of the initial enzyme specific activity was retained after immobilization in the foam. Factors affecting the rate of paraoxon degradation include foam hydrophobicity, the degree of mixing applied to initiate polymerization, and foam pretreatment prior to use in substrate hydrolysis. The storage stability of the foam is significant, with phosphotriesterase-foam activity profiles exhibiting a three month half-life. Foams are currently being developed for biocatalytic air filtering, in which gaseous substrates will be simultaneously adsorbed and degraded by the immobilized enzyme system. © 1996 John Wiley & Sons, Inc.
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    Continuous cultures limited by a gaseous substrate: Development of a simple, unstructured mathematical model and experimental verification with Methanobacterium thermoautotrophicum (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 51 (1996), S. 645-658 
    Details
    ISSN: 0006-3592
    Keywords: Methanobacterium thermoautotrophicum ; gaseous substrate limitation ; continuous culture ; mathematical modeling ; amperometric measurement of dissolved H2 concentration ; reaction calorimetry ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: This article presents a simple, unstructured mathematical model describing microbial growth in continuous culture limited by a gaseous substrate. The model predicts constant gas conversion rates and a decreasing biomass concentration with increasing dilution rate. It has been found that the parameters influencing growth are primarily the gas transfer rate and the dilution rate. Furthermore, it is shown that, for correct simulation of growth, the influence of gaseous substrate consumption on the effective gas flow through the system has to be taken into account.Continuous cultures of Methanobacterium thermoautotrophicum were performed at three different gassing rates. In addition to the measurement of the rates of biomass production, product formation, and substrate consumption, microbial heat dissipation was assessed using a reaction calorimeter. For the on-line measurement of the concentration of the growth-limiting substrate, H2, a specially developed probe has been used. Experimental data from continuous cultures were in good agreement with the model simulations. An increase in gassing rate enhanced gaseous substrate consumption and methane production rates. However, the biomass yield as well as the specific conversion rates remained constant, irrespective of the gassing rate. It was found that growth performance in continuous culture limited by a gaseous substrate is substantially different from “classic” continuous culture in which the limiting substrate is provided by the liquid feed. In this report, the differences between both continuous culture systems are discussed.
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    Plasmid stability of recombinant Pseudomonas sp. B13 FR1 pFRC20P in continuous culture (1998)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 57 (1998), S. 62-70 
    Details
    ISSN: 0006-3592
    Keywords: plasmid stability ; recombinant microorganism ; continuous culture ; Pseudomonas sp. B13 FR1 pFRC20P ; degradation of aromatic compounds ; chlorobenzoate ; methylbenzoate ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Plasmid stability of recombinant Pseudomonas sp. B13 FR1 pFRC20P, a strain capable of mineralizing 3- and 4-chlorobenzoate and 4-methylbenzoate, was investigated in continuous culture. The hybrid cosmid pFRC20P enables the strain to mineralize 4-methylbenzoate. Rapid plasmid loss was observed under nonselective conditions using 3-chlorobenzoate as the substrate. Plasmid stability decreased with increasing dilution rate. Despite the growth advantage of the generated plasmid free cells a total depletion of plasmid bearing cells was not observed. After approximately 50 generations the fraction of plasmid bearing cells reached a constant level of 10%, which was stably maintained during the next 25 generations. Cells from this stage were used to inoculate a new culture that resulted in a stable level of 50% plasmid bearing cells. By a temporary substrate change to selective conditions (4-methylbenzoate), this level could be further increased to 70%. Literature models on plasmid stability could not be applied to describe the experimental data. Therefore, a new but unstructured model was developed to describe the experimental results. The model is based on the existence of three subpopulations: a plasmid free one, an original plasmid bearing one with a growth disadvantage compared to plasmid free cells, and a second plasmid bearing subpopulation with increased stability that is generated from the original one and has a growth rate comparable to the plasmid free cells. © 1998 John Wiley & Sons, Inc. Biotechnol Bioeng 57: 62-70, 1998.
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    The use of confocal scanning laser microscopy and other tools to characterize Escherichia coli in a high-cell-density synthetic biofilm (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 52 (1996), S. 340-356 
    Details
    ISSN: 0006-3592
    Keywords: laser microscopy, confocal scanning ; Escherichia coli ; biofilms ; immobilization ; confocal scanning laser microscopy ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Properties of a novel, synthetic biofilm were examined by using confocal scanning laser microscopy (CSLM) in combination with fluorescent probes and by investigating total protein content and specific β-galactosidase activity during various steps of the biofilm preparation. Viable, but nongrowing Escherichia coli were entrapped in 10- to 80-μm-thick multilayer films, where a copolymer of acrylic and vinyl acetate was the immobilization matrix. Cell viability and distribution within the films were evaluated by developing a protocol to stain the bacteria with fluorescein isothiocyanate and propidium iodide, thereby labeling all cells green and dead cells red, respectively. Confocal microscopy facilitated viewing samples in the XY and XZ planes, and image analysis enabled counting of the cells. These experiments showed that the initial viability of the entrapped bacteria was 85% to 90%, cell distribution was uniform in the XY plane and cell number increased with increasing depth into the film. Specific β-galactosidase assays developed here allowed comparison of the induction of lacZin suspended and immobilized cells. These experiments demonstrated that rehydration was an important step in biofilm preparation, and E. coli cast into synthetic biofilms with cell layers of at least 20 to 35 μm in thickness had gene induction characteristics similar to suspended cells. © 1996 John Wiley & Sons, Inc.
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    Immobilization of manganese peroxidase from Lentinula edodes and its biocatalytic generation of MnIII-chelate as a chemical oxidant of chlorophenols (1998)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 60 (1998), S. 204-215 
    Details
    ISSN: 0006-3592
    Keywords: immobilization ; white-rot fungi ; Lentinula edodes ; manganese peroxidase ; Mn3+ ; azlactone ; chlorophenol ; EEDQ ; biocatalyst ; bioremediation ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Manganese peroxidase (MnP) purified from commercial cultures of Lentinula edodes was covalently immobilized through its carboxyl groups using an azlactone-functional copolymer derivatized with ethylenediamine and 2-ethoxy-1-ethoxycarbonyl-1,2-dihydroquinoline (EEDQ) as a coupling reagent. The tethered enzyme was employed in a two-stage immobilized MnP bioreactor for catalytic generation of chelated MnIII and subsequent oxidation of chlorophenols. Manganese peroxidase immobilized in the enzyme reactor (reactor 1) produced MnIII-chelate, which was pumped into another chemical reaction vessel (reactor 2) containing the organopollutant. Reactor 1-generated MnIII-chelates oxidized 2,4-dichlorophenol and 2,4,6-trichlorophenol in reactor 2, demonstrating a two-stage enzyme and chemical system. H2O2 and oxalate chelator concentrations were varied to optimize the immobilized MnP's oxidation of MnII to MnIII. Oxidation of 1.0 mM MnII to MnIII was initially measured at 78% efficiency under optimized conditions. After 24 h of continuous operation under optimized reaction conditions, the reactor still oxidized 1.0 mM MnII to MnIII with ∼69% efficiency, corresponding to 88% of the initial MnP activity. © 1998 John Wiley & Sons, Inc. Biotechnol Bioeng 60: 204-215, 1998.
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    Trehalase immobilization on aminopropyl glass for analytical use (1997)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 54 (1997), S. 33-39 
    Details
    ISSN: 0006-3592
    Keywords: trehalase ; trehalose ; immobilization ; aminopropyl glass ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Trehalase is the enzyme which hydrolyzes the disaccharide trehalose into two α-D-glucose molecules. In this article, we present the immobilization of trehalase on aminopropyl glass particles. The enzyme was extracted from Escherichia coli Mph2, a strain harboring the pTRE11 plasmid, which contains the trehalase gene. The partially purified enzyme had a specific activity of 356 U/mg and could be used for quantifying trehalose in the presence of sucrose, maltose, lactose, starch, and glycogen. Partially purified trehalase was immobilized by covalent coupling with retention of its catalytic activity. The support chosen for the majority of the experiments reported was aminopropyl glass, although spherisorb-5NH2 and chitin were also tested. The immobilized enzyme was assayed continuously for 40 h, at pH 6.0 and 30°C, and no release of enzyme molecules was detected during this procedure. The best condition found for storing the enzyme-support complex was at 4°C in the presence of 25 mM sodium maleate, containing 7 mM β-mercaptoethanol, 1 mM ethylenediaminetetraacetic acid (EDTA), and 50% glycerol. The enzyme under these conditions was stable, retaining approximately 100% of its initial activity for at least 28 days. The immobilized enzyme can be employed to detect trehalose molecules in micromolar concentration. The optimum pH value found was 4.5 and the Km app. 4.9 × 10-3 M trehalose at pH 4.6 and 30°C, with Vmax of 5.88 μmol glucose · min.-1, as calculated by a Lineweaver-Burk plot. © 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 54: 33-39, 1997.
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    The effect of dissolved oxygen on the metabolic profile of a murine hybridoma grown in serum-free medium in continuous culture (1997)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 54 (1997), S. 153-164 
    Details
    ISSN: 0006-3592
    Keywords: hybridoma ; oxygen ; serum-free medium ; continuous culture ; antioxidant ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The murine B-lymphocyte hybridoma, CC9C10 was grown at steady state under serum-free conditions in continuous culture at dissolved oxygen (DO) concentrations in the range of 10% to 150% of air saturation. Cells could be maintained with this range at high viability in a steady state at a dilution rate of 1 d-1, although with lower cell concentrations at higher DO. A higher specific antibody production measured at higher DO was matched by a decrease in the viable cell concentration at steady state, so that the volumetric antibody titre was not changed significantly. An attempt to grow cells at 250% of air saturation was unsuccessful but the cells recovered to normal growth once the DO was decreased.There was a requirement for cellular adaptation at each step-wise increase in dissolved oxygen. Adaptation to a DO of 100% was associated with an increase in the specific activities of glutathione peroxidase (×18), glutathione S-transferase (×11) and superoxide dismutase (×6) which are all known antioxidant enzymes. At DO above 100%, the activities of GPX and GST decreased possibly as a result of inactivation by reactive oxygen radicals.The increase in dissolved oxygen concentration caused changes in energy metabolism. The specific rate of glucose uptake increased at higher dissolved oxygen concentrations with a higher proportion of glucose metabolized anaerobically. Short-term radioactive assays showed that the relative flux of glucose through glycolysis and the pentose phosphate pathway increased whereas the flux through the tricarboxylic acid cycle decreased at high DO. Although the specific glutamine utilization rate increased at higher DO, there was no evidence for a change in the pattern of metabolism. This indicates a possible blockage of glycolytic metabolites into the TCA cycle, and is compatible with a previous suggestion that pyruvate dehydrogenase is inhibited by high oxygen concentrations.Analysis of the oxygen uptake rate of cell suspensions at steady state under all conditions showed a pronounced Crabtree effect which was manifest by a decrease (up to 40%) in oxygen consumption on addition of glucose. This indicates that the degree of aerobic metabolism in these cultures is highly sensitive to the glucose concentration. © 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 54: 153-164, 1997.
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    Removal of chlorophenols from wastewater by immobilized horseradish peroxidase (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 51 (1996), S. 126-130 
    Details
    ISSN: 0006-3592
    Keywords: chlorophenol ; peroxidase ; immobilization ; magnetite ; immobilized enzyme ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Immobilization of horseradish peroxidase on magnetite and removal of chlorophenols using immobilized enzyme were investigated. Immobilization by physical adsorption on magnetite was much more effective than that by the crosslinking method, and the enzyme was found to be immobilized at 100% of retained activity. In addition, it was discovered that horseradish peroxidase was selectively adsorbed on magnetite, and the immobilization resulted in a 20-fold purification rate for crude enzyme. When immobilized peroxidase was used to treat a solution containing various chlorophenols, p-chlorophenol, 2,4-dichlorophenol, 2,4,5-trichlorophenol, 2,4,6-trichlorophenol, 2,3,4,6-tetrachlorophenol, and pentachlorophenol, each chlorophenol was almost 100% removed, and also the removal of total organic carbon (TOC) and adsorbable organic halogen (AOX) reached more than 90%, respectively. However, in the case of soluble peroxidase, complete removal of each chlorophenol could not be attained, and in particular, the removal of 2,4,5-trichlorophenol was the lowest, with a removal rate of only 36%. © 1996 John Wiley & Sons, Inc.
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    Immobilization of trypsin onto “molded” macroporous poly(glycidyl methacrylate-co-ethylene dimethacrylate) rods and use of the conjugates as bioreactors and for affinity chromatography (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 49 (1996), S. 355-363 
    Details
    ISSN: 0006-3592
    Keywords: trypsin ; immobilization ; molded support ; poly(glycidyl methacrylate-co-ethylene dimethacrylate) ; porous materials ; affinity chromatography ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Trypsin immobilization onto continuous “molded” rods of porous poly(glycidyl methacrylate-co-ethylene dimethacrylate) and some applications of the conjugate have been studied. The rods polymerized within a tubular mold (chromatographic column), were treated in situ with ethylenediamine, activated with glutaraldehyde and finally modified with trypsin. The performance of the trypsin-modified rods was evaluated and compared to that of poly(glycidyl methacrylate-co-ethylene dimethacrylate) beads, modified with the same enzyme. Overall the enzyme-modified rods performed substantially better than the corresponding beads. In particular, the performance of the molded supports as enzymatic reactors or as chromatographic media benefits greatly from the enhanced mass transfer that is characteristic of the molded rod at high flow rates. © 1996 John Wiley & Sons, Inc.
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    Influence of the degree and mode of light limitation on growth characteristics of the Rhodobacter capsulatus continuous cultures (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 51 (1996), S. 605-612 
    Details
    ISSN: 0006-3592
    Keywords: phototrophic bacteria ; Rhodobacter capsulatus ; continuous culture ; light limitation ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The influence of the degree and mode of light limitation on growth characteristics of turbidostat cultures of Rhodobacter capsulatus was investigated using mass and energy balance regularities. Light limitation was achieved by increasing the steady-state biomass concentration at constant incident light intensity (∼100 W/m2) or by decreasing the incident light intensity at constant steady-state biomass concentration (∼500 mg of dry biomass/L). It was shown that under conditions of light limitation of Rh. capsulatus, the content of P and N in the biomass as well as the biomass degree of reduction were determined by the growth rate of the cultures. The energetic yield of biomass of Rh. capsulatus and total bacteriochlorophyll a content increased when light limitation increased. These parameters were higher in the cultures, in which light limitation was achieved by lowering the incident light intensity at low biomass concentration. This seems to be due to different distribution of light within the photobioreactor when dissimilar modes of light limitation were used.
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    Kinetic evaluation of biotransformation of benzaldehyde to L-phenylacetylcarbinol by immobilized pyruvate decarboxylase from Candida utilis (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 49 (1996), S. 429-436 
    Details
    ISSN: 0006-3592
    Keywords: biotransformation ; L-phenylacetylcarbinol ; immobilization ; pyruvate decarboxylase ; Candida utilis ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Biotransformation of benzaldehyde to L-phenylacetylcarbinol (L-PAC) as a key intermediate for L-ephedrine has been evaluated using immobilized pyruvate decarboxylase (PDC) from Candida utilis. PDC immobilized in spherical polyacrylamide beads was found to have a longer half-life compared with free enzyme. In a batch process, the immobilized PDC generally produced lower L-PAC than free enzyme at the same concentrations of substrates due to increased by-products acetaldehyde and acetoin and reduced benzaldehyde uptake. With immobilized PDC, L-PAC formation occurred at higher benzaldehyde concentrations (up to 300 mM) with the highest L-PAC concentration being 181 mM (27.1 g/L). For a continuous process, when 50 mM benzaldehyde and 100 mM sodium pyruvate were fed into a packed-bed reactor at 4°C and pH 6.5, a productivity of 3.7 mM/h (0.56 g/L · h) L-PAC was obtained at an average concentration of 30 mM (4.5 g/L). The half-life of immobilized PDC reactor was 32 days. © 1996 John Wiley & Sons, Inc.
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    Efficient immobilization of lipases by entrapment in hydrophobic sol-gel materials (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 49 (1996), S. 527-534 
    Details
    ISSN: 0006-3592
    Keywords: lipase ; immobilization ; sol-gel materials ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The commercial application of lipases as biocatalysts for organic synthesis requires simple but efficient methods to immobilize the enzyme, yielding highly stable and active biocatalysts which are easy to recover. In this study, we present a novel method to achieve lipase immobilization by entrapment in chemically inert hydrophobic silica gels which are prepared by hydrolysis of alkyl-substituted silanes in the presence of the enzyme. A typical immobilization procedure uses: an aqueous solution of lipase; sodium fluoride as a catalyst; and additives like polyvinyl alcohol or proteins and alkoxysilane derivatives like RSi-(OMe)3 with R = alkyl, aryl, or alkoxy as gel precursors. The effect of various immobilization parameters like stoichiometric ratio of water, silane, type and amount of additive, type and amount of catalyst, and type of silane has been carefully studied. The new method is applicable for a wide variety of lipases, yielding immobilized lipases with esterification activities enhanced by a factor of up to 88, compared to the commercial enzyme powders under identical conditions. Studies on the stability of sol-gel immobilized lipases under reaction conditions or storage (dry, in aqueous or organic medium) revealed an excellent retention of enzymatic activity. The possible reasons for the increased enzyme activities are discussed. © 1996 John Wiley & Sons, Inc.
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    The application of membrane biological reactors for the treatment of wastewaters (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 49 (1996), S. 601-610 
    Details
    ISSN: 0006-3592
    Keywords: aerobic ; anaerobic ; biomass separation ; bioreactor ; bubbleless ; oxygen mass transfer ; extraction of organic pollutants ; membrane ; wastewaters ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Combining membrane technology with biological reactors for the treatment of municipal and industrial wastewaters has led to the development of three generic membrane processes within bioreactors: for separation and recycle of solids; for bubbleless aeration of the bioreactor; and for extraction of priority organic pollutants from hostile industrial wastewaters. Commercial aerobic and anaerobic membrane separation bioreactors already provide a small footprint alternative to conventional biological treatment methods, producing a high-quality effluent at high organic loading rates. Both the bubbleless aeration and extractive membrane bioreactors are in the development stages. The former uses gas-permeable membranes to improve the mass transfer of oxygen to the bioreactor by providing bubbleless oxygen. By using a silicone membrane process, extractive membrane bioreactors transfer organic pollutants from chemically hostile wastewaters to a nutrient medium for subsequent biodegradation. All three membrane bioreactor (MBR) processes are comparatively and critically reviewed. © 1996 John Wiley & Sons, Inc.
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    Application of light-emitting diodes in bioreactors: Flashing light effects and energy economy in algal culture (Chlorella pyrenoidosa) (1996)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 50 (1996), S. 98-107 
    Details
    ISSN: 0006-3592
    Keywords: algal culture ; bioreactor ; bioregenerative system ; energy economy ; light-emitting diode (LED) ; microsecond pulse modulation ; Chlorella pyrenoidosa ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Light-emitting diodes (LEDs) were used as the sole light source in continuous culture of the green alga Chlorella pyrenoidosa. The LEDs applied show a peak emission at 659 nm with a half-power bandwidth of 30 nm. Selection of this wavelength range, which is optimal for excitation of chlorophylls a and b in their “red” absorption bands makes all photons emitted potentially suitable for photosynthesis. No need for additional supply of blue light was found. A standardized panel with 2 LEDs cm-2 fully covered one side of the culture vessel. At standard voltage in continuous operation the light output of the diode panel appeared more than sufficient to reach maximal growth. Flash operation (5-μs pulse duration) enables potential use of higher operating voltages which may render up to three times more light output. Flat airlift fermentor-type continuous culture devices were used to estimate steady state growth rates of Chlorella pyrenoidosa as a function of the light flux (μmol photons · m-2 · s-1) and the flashing frequency of the light-emitting diodes (which determines the duration of the dark “off” time between the 5-μs “on” pulses). At the fixed voltage and turbidostat setting applied a 20-kHz frequency, which equals dark periods of 45 μs, still permitted the maximum growth rate to become nearly reached. Lower frequencies fell short of sustaining the maximal growth rate. However, the light flux decrease resulting from lowering of the flash frequency appeared to reduce the observed growth rates less than in the case of a similar flux decrease with light originating from LEDs in continuous operation. Flash application also showed reduction of the quantum requirement for oxygen evolution at defined frequencies. The frequency domain of interest was between 2 and 14 kHz. LEDs may open interesting new perspectives for studies on optimization of mixing in mass algal culture via the possibility of separation of interests in the role of modulation on light energy conversion and saturation of nutrient supply. Use of flashing LEDs in indoor algal culture yielded a major gain in energy economy in comparison to luminescent light sources. © 1996 John Wiley & Sons, Inc.
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    Thermophilic sulphate and sulphite reduction in lab-scale gas-lift reactors using H2 and CO2 as energy and carbon source (1997)
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 55 (1997), S. 807-814 
    Details
    ISSN: 0006-3592
    Keywords: sulphate reduction ; sulphite reduction ; biofilm ; immobilization ; gas-lift reactor ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Feasibility of thermophilic (55°C) sulphate and sulphite reduction with H2 and CO2 gas-mixtures was studied in gas-lift reactors, which contained pumice particles as carrier material. Particular attention was paid to biomass retention and the competition between hydrogenotrophic sulphate-reducers and other hydrogenotrophic thermophiles. A model medium with defined mineral nutrients was used.The results of the experiments clearly demonstrate that sulphate conversion rates up to 7.5 g SO42-/L per day can be achieved. With sulphite, a reduction rate of 3.7 g S/L per day was obtained, which equals a sulphate conversion rate of 11.1 g SO42-/L per day. Under the applied conditions, a strong competition for hydrogen between hydrogenotrophic sulphate-reducers, tentatively designated as Desulfotomaculum sp., and hydrogenotrophic methanogens was observed. The outcome of the competition could not be predicted. Growth of the mixed culture was totally inhibited at an H2S concentration of 250 mg/L. Poor attachment of sulphate-reducing bacteria was observed in all experiments. The biomass concentration did not exceed 1.2 g/L, despite the presence of 50 g/L of pumice. The reason for this phenomenon remains to be understood. © 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 55: 807-814, 1997.
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