bluuebunny/crossref_metadata_2025_split huggingface dataset:
Microencapsulation selection for isolation of yeast mutants with increased secretion of Aspergillus awamori glucoamylase
Authors: Wittrup K. D. , Weber A. L. , Tsai P. 
Published: December 2004

AbstractWe have developed a microencapsulation selection method which allows the rapid and quantitative screening of >106 yeast cells for enhanced secretion of Aspergillus awamori glucoamylase. The method provides a 400‐fold single‐pass enrichment for high‐secreting mutants, and can be straightforwardly adapted for application to growth‐based selection schemes with other microorganisms and enzymes. © 1993 John Wiley & Sons, Inc.
Monoclonal antibody productivity and the metabolic pattern of perfusion cultures under varying oxygen tensions
Authors: Shi Yuan , Ryu Dewey D. Y. , Park Sun Ho 
Published: December 2004

AbstractThe metabolic pattern and cell culture kinetics of high‐cell‐density perfusion cultures were compared under two different oxygen transfer conditions: oxygen limiting and not limiting. When oxygen was a limiting factor during perfusion culture, both specific glucose uptake and lactate production rates increased, compared to non‐oxygen‐limited condition, by about 60% and 30%, respectively. The specific glutamine uptake rate under oxygen‐limited conditions was almost 4.0 times higher than that under non‐oxygen‐limited conditions. The activity of lactate dehydrogenase (LDH) released into the medium by the dead cells can be used as an indicator for the metabolic and physiological conditions related to oxygen limitation. There was a 3.2 times higher specific rate of LDH activity released by dead cells in oxygen‐limited cultures than those in non‐oxygen‐limited cultures. The specific production rate of monoclonal antibody was not significantly affected by the oxygen transfer conditions during the rapid cell growth period, but it rapidly increased toward the end of perfusion cultures. The higher perfusion rate may have limited further cell growth during high‐cell‐density perfusion culture, because cell damage was caused by the hydrodynamic shear within a hollow fiber microfiltration cartridge installed to withdraw the spent medium and the waste metabolites. © 1993 John Wiley & Sons, Inc.
Competition between two microbial populations in a sequencing fed‐batch reactor: Theory, experimental verification, and implications for waste treatment applications
Authors: Dikshitulu S. , Baltzis B. C. , Lewandowski G. A. , Pavlou S. 
Published: December 2004

AbstractCompetition between two microbial populations for a single pollutant (phenol) was studied in a sequencing fed‐batch reactor (SFBR). A mathematical model describing this system was developed and tested experimentally. It is based on specific growth rate expressions revealed from pure culture batch experiments. The species employed were Pseudomonas putida (ATCC 17514) and Pseudomonas resinovorans (ATCC 14235). It was found that both species biodegrade phenol following inhibitory kinetics which can be described by Andrews' expression. The model predicts that the dynamics of a SFBR, and the kinetics of biodegradation, result in a complex set of operating regimes in which neither species, only one species, or both species can survive at steady cycle. The model also predicts the existence of multiple outcomes, achievable from different start‐up conditions, in some domains of the operating parameter space. Experimental results confirmed the model predictions. There was excellent agreement between predicted and measured concentrations of phenol, total biomass, and the biomass of each individual species. This study shows how serious discrepancies can arise in scale‐up of biodegradation data if population dynamics are not taken into account. It also further confirms experimentally the theory of microbial competition in periodically forced bioreactors. © 1993 John Wiley & Sons, Inc.
Enhanced production of α‐amylase in fed‐batch cultures of Bacillus subtilis TN106[pAT5]
Authors: Lee Jeewon , Parulekar Satish J. 
Published: December 2004

AbstractGrowth of Bacillus subtilis TN106[pAT5] and synthesis of plasmid‐encoded protein (α‐amylase) are investigated in batch, continuous, and fed‐batch cultures using a defined medium containing glucose and/or starch as the carbohydrate source. The batch culture studies reveal that reduced availability of arginine hampers growth of recombinant cells (which lack an arginine synthesis gene) but promotes production of α‐amylase and substitution of glucose by starch as the carbohydrate source leads to slower growth of recombinant cells and increased production of α‐amylase per unit cell mass. Retention of recombinant cells over prolonged periods in continuous cultures is not possible without continuous application of antibiotic selection pressure owing to segregational plasmid instability. Fed‐batch experiments with constant volumetric feed rate demonstrate that α‐amylase production is enhanced at lower feed concentration of starch (sole carbohydrate source) and lower volumetric feed rate. Such slow addition of starch is however not conducive for growth of recombinant cells. The expression of the thermostable α‐amylase gene carried on the recombinant plasmid pAT5 (derived from a plasmid isolated from a thermophilic bacterium) is promoted at higher temperatures, while growth of recombinant cells is depressed. In all batch and fed‐batch experiments, production of α‐amylase is observed to be inversely related to growth of recombinant cells. The efficacy of two‐stage bioreactor operations, with growth of recombinant cells being promoted in the first stage and α‐amylase production in the second stage, in attaining increased bulk α‐amylase activity is demonstrated. © 1993 John Wiley & Sons, Inc.
Repeated fed‐batch culture of hybridoma cells in nutrient‐fortified high‐density medium
Authors: Jo Eui‐Cheol , Park Hae‐Joon , Kim Dong‐II , Moon Hong Mo 
Published: December 2004

AbstractLong‐term high‐density cultivation of the hybridoma 2c3.1 was successfully carried out in a repeated fed‐batch mode using high‐density media that were constructed to meet in vitro cell growth limitations. The high‐density culture was possible in a range of 0.5 ∼ 1.0 × 107 cells/mL in MBRI 40‐02 medium for over 2500 h by the repeated supplementation of the most fortified medium, MBRI 40‐03, and consequently, distinct enhancement of MAb production was achieved. MAb concentrations were maintained around 1 g/L for about 1000 h of the process and the maximum MAb concentration was around 1.56 g/L. The result supported strongly the fact that the nutritional fortification was the most critical factor for high‐density cell culture in vitro. The mean chromosome number of the hybridoma 2c3.1 was maintained stably for about 1500 h, whereas gradual loss of the MAb activity was apparent during the long‐term cultivation. © 1993 John Wiley & Sons, Inc.
Adsorption–desorption of recombinant hepatitis B surface antigen (r‐HBsAg) from P. pastoris on a diatomaceous earth matrix: Optimization of parameters for purification
Authors: Agraz Alberto , Quiñones Yair , Expósito Néstor , Breña Fidel , Madruga Joel , Pentón Eduardo , Herrera Luis 
Published: December 2004

AbstractRecombinant hepatitis B surface antigen (r‐HBsAg) produced in yeast is adsorbed on a diatomaceous earth matrix for purification purposes. A pH dependence in the adsorption–elution behavior was found. The capacity of celite (Hyflo Super Cei) for adsorbing r‐HBsAg increased with decreasing pH. Nonspecific proteins were also adsorbed, but a low pH dependence was found. Elution from the matrix was performed using a basic pH buffer, in which r‐HBsAg is more specifically adsorbed/desorbed than contaminant proteins, permitting the purification of the r‐HBsAg. A pH of 4.0 was used for adsorption and pH 8.2 was used for desorption. The described protocol allows a purification factor between three‐ and fivefold with respect to contaminant proteins and sixfold with respect to contaminant DNA. Finally, the adsorption step was successfully scaled‐up for production purposes. © 1993 John Wiley & Sons, Inc.
Scale‐up on basis of structured mixing models: A new concept
Authors: Mayr B. , Moser A. , Nagy E. , Horvat P. 
Published: December 2004

AbstractA new scale‐up concept based upon mixing models for bioreactors equipped with Rushton turbines using the tanks‐in‐series concept is presented. The physical mixing model includes four adjustable parameters, i.e., radial and axial circulation time, number of ideally mixed elements in one cascade, and the volume of the ideally mixed turbine region. The values of the model parameters were adjusted with the application of a modified Monte‐Carlo optimization method, which fitted the simulated response function to the experimental curve. The number of cascade elements turned out to be constant (N = 4). The model parameter radial circulation time is in good agreement with the one obtained by the pumping capacity. In case of remaining parameters a first or second order formal equation was developed, including four operational parameters (stirring and aeration intensity, scale, viscosity). This concept can be extended to several other types of bioreactors as well, and it seems to be a suitable tool to compare the bioprocess performance of different types of bioreactors. © 1994 John Wiley & Sons, Inc.
Continuous culture of Methanococcus jannaschii, an extremely thermophilic methanogen
Authors: Tsao Jia‐Huey , Kaneshiro Stacey M. , Yu Shu‐San , Clark Douglas S. 
Published: December 2004

AbstractMethanococcus jannaschii, an extremely thermophilic methanogen isolated from a deep‐sea hydrothermal vent, was grown at 80°C in continuous culture on a mineral salts medium gassed with H2 and CO2 at three different flow rates. The maximum specific growth rate was 0.56 h−1, and the maximum specific methane productivity was 0.32 (mol g−1 h−1). Uncoupling of growth and methane production was evidenced by an increase in teh non‐growth‐associated rate of methane formation, β, with increasing gaseous input. The specific hydrogenase activity exhibited growth‐assiciated behaviour at low growth rates, but showed no dependence on growth at higher growth rates. The growth dependence of hydrogenase activity is consistent with the pressure dependence of hydrogenase activity measured in previous experiments. In contrast, the specific protease activity was independent of the growth rate over the entire range of dilution rates studied. © 1994 John Wiley & Sons, Inc.
Predictions for oxygen supply control to enhance population stability of engineered production strains
Authors: Varma Amit , Palsson Bernhard O. 
Published: December 2004

AbstractThe simultaneous growth and product formation in a microbial culture is an important feature of several laboratory, industrial, and environmental bioprocesses. Metabolic burden associated with product formation in these bioprocesses may lead to growth advantage of a nonproducing mutant leading to a loss of the producing population over time. A simple population dynamics model demonstrates the extreme sensitivity of population stability to the engineered productivity of a strain. Here we use flux balance analysis to estimate the effects of the metabolic burden associated with product secretion on optimal growth rates. Comparing the optimal growth rates of the producing and nonproducing strains under a given processing condition allows us to predict the population stability. In order to increase stability of an engineered strain, we determine processing conditions that simultaneously maximize the growth rate of the producing population while minimizing the growth rate of a nonproducing population. Using valine, tryptophan, and lysine production as specific examples, we demonstrate that although an appropriate choice of oxygenation may increase culture longevity more than twofold, total production as governed by economic criterion can be increased by several orders of magnitude. Choice of optimal nutrient and oxygen supply rates to enhance stability is important both for strain screening as well as for culture of engineered strains. Appropriate design of the culture environment can thus be used to enhance the productivity of bioprocesses that use engineered production strains. © 1994 John Wiley & Sons, Inc.
Multifunctional poly(ethylene glycol) semi‐interpenetrating polymer networks as highly selective adhesive substrates for bioadhesive peptide grafting
Authors: Drumheller Paul D. , Elbert Donald L. , Hubbell Jeffrey A. 
Published: December 2004

AbstractNovel artificial extracellular matrices were synthesized in the form of semi‐interpenetrating polymer networks containing copolymers of poly(ethylene glycol) and acrylic acid (PEG–co‐AA) grafted with synthetic bioadhesive peptides onto exposed carboxylic acid moieties. These substrates were very resistant to cell adhesion, but when they were grafted with adhesive peptides they were highly biospecific in their ability to support cell adhesion. Extensive preadsorption of adhesive proteins or peptides did not render these materials cell adhesive; yet covalent grafting of adhesive peptides did render these materials highly cell adhesive even in the absence of serum proteins. Polymer networks containing immobilized PEG–co‐AA were grafted with peptides at densities of 475 ± 40 pmol/cm2. Polymer networks containing immobilized PEG‐co‐AA N‐terminally grafted with GRGDS supported cell adhesion efficiencies of 42 ± 4% 4 h after seeding and became confluent after 12 h. These cells displayed cell spreading and cytoskeletal grafted with inactive control peptides (GRDGS, GRGES, or no peptide) supported cell adhesion efficiencies of 0 ± 0%, even when challenged with high seeding densities (to 100,000 cell/cm2) over 14 days. These polymer networks are suitable substrates to investigate in vitro cell‐surface interactions in the presence of serum proteins without nonspecific protein adsorption adhesion signals other than those immobilized for study.
Cerebral neurons, skeletal myoblasts, and cardiac muscle cells cultured on macroporous beads
Authors: Shahar A. , Reuveny S. , David Y. , Budu C. , Shainberg A. 
Published: December 2004

AbstractBiodegradable macroporous spherical microcarriers (MCs) offer a suitable substrate for adhesion, growth, and differentiation of cerebral neurons, skeletal myoblasts, and cardiac cells. The cavernous structure of these gelatin beads supplies a large surface and a tridimensional habitat for cell propagation. Within hours from their seeding, all three cell types became firmly attached to the MCs, forming cell–MC aggregates, which remained floating in the medium. Neuronal aggregates were composed mainly of single or groups of perikarya, and their sprouted nerve processes formed a ramified network. In skeletal muscle aggregates, fusion of myoblasts into myotubes occured within 5 days. The myotubes, arranged in bundles having the same orientation, became striated and the whole aggregate, or parts of it, contracted spontaneously. Cardiac cells divided in the aggregate to form one or more layers of flat cells, which exhibited single microvilli. The cells were interconnected at uneven intervals and the whole aggregate contracted actively. Morhological and biochemical analysis of sampled aggregates ensured that cells reached the proper stage of maturation and could be used for either physiological and pharmacological studies or implantation into injured or dystrophic tissue.
Baculovirus expression system scaleup by perfusion of high‐density Sf‐9 cell cultures
Authors: Caron Antoine W. , Tom Rosanne L. , Kamen Amine A. , Massie Bernard 
Published: December 2004

AbstractA perfusion system based on a 4‐L stirred tank bioreactor and a custom‐designed tangential (cross‐flow) filter was assembled to realize a scaleup of the Baculovirus Expression Vector System (BEVS). When perfused with 1 to 1.5 vol/day, Spodoptera frugiperda (Sf‐9) insect cell cultures grew from 4 × 106 to 15 × 106 cells/mL over 3 to 4 days. The possibility of maintaining high specific production of recombinant VP6 protein (from bovine rotavirus) after baculovirus infection of the high‐density cultures was then assessed. The process consisted of a growth phase in TNMFH + 10% FBS, followed by infection with Bac‐BRV6L recombinant baculovirus and a shift to a low‐serum (0 to 1%) medium for perfusion during the production phase. Multiple runs were executed, each including a battery of shaker flask controls at various cell densities and serum concentrations. On average, specific rVP6 production in the bioreactor amounted to 76% of that found in 20‐mL shaker cultures simulatingthe bioreactor's high cell density, low serum concentration, and medium renewal rate. Mechanical stress generated by cell/medium separation in theperfusion process reduced cell growth rate but had minimal effect on rVP6production. Our results also indicated that serum concentration during the infection phase affected the rVP6 specific production in a cell density–dependent fashion. Although the feasibility of the cell density scale up was demonstrated, optimization is still needed to achieve a truly cost‐effective process.
Synthesis of aspartame precursor with an immobilized thermolysin in tert‐amyl alcohol
Authors: Nagayasu Takeshi , Miyanaga Masamitsu , Tanaka Takaaki , Sakiyama Takaharu , Nakanishi Kazuhiro 
Published: December 2004

AbstractN‐(Benzyloxycarbonyl)‐L‐aspartyl‐L‐phenylalanine methyl ester (Z‐AspPheOMe), a precursor of the synthetic sweetner asparatame, was synthesized from N‐(benzyloxycarbolyl)‐L‐aspartic acid (Z‐Asp) and L‐phenylalanine methyl ester (PheOMe) with an immobilized thermolysin in various organic solvents. We found that in tert‐amyl alcohol containing a small amount of water the immobilized enzyme showed a high activity comparble to that in ethyl acetate with quite a high stability. The immobilized enzyme was fully stable up to 70°C in tert‐amyl alcohol in the absence of the subatrate, and up to 50°C in the presence of the substrate. The high stability in the presence of the substrate was found due to the fact that the release of calcium ions, the stabilizing factor of thermolysin, is suppressed.The substrate concentration dependence of the initial synthetic rate with the immobilized enzyme was quite different from that with the free enzyme in the biphasic system, in contrast to that in ethyl acetate. Finally, Z‐AspPheOMe was continuously synthesized in a column reactor using 200 mM PheOMe and 120 mM Z‐Asp as the substrate for over 300 h at 45°C and a space velocity of 1 h−1 without any loss of acivity. © 1994 John Wiley & Sons, Inc.
Stoichiometric analysis of animal cell growth and its application in medium design
Authors: Xie Liangzhi , Wang Daniel I. C. 
Published: December 2004

AbstractAnimal cell cultivation in vitro has been studied for more than 40 years. However, the culture medium composition has not been designed on the basis of the stoichiometric nutritional demands for animal cell growth. In this article, a model was developed to study the stoichiometric demands for nutrients (including glucose, 20 amino acids, and 10 vitamins)for the synthesis of cell mass and product. The coefficients for these nutrients in the stoichiometric equation governing animal cell growth were determined based on cell composition. In addition, a detailed analysis of the nutrients′ roles in the synthesis of cell mass and product was also performed. Applications of the stoichiometric analysis in animal cell cultivation, such as culture medium design, supplemental medium formulation, and feeding strategy will also be discussed. The stoichiometric analysis can be potentially employed to analyze results from animal cell cultures, to improve the performance of culture processes, and to design new process rationally. It can also help to provide a better understanding of animal cell metabolism. Simplifications on the cellular energy metabolism were made in order to simplify the model and to provide the preliminary bases to test the process performance. However, this could introduce inaccuracies for the model and results in errors in the calculations of glucose and glutamine concentrations when employed in medium design. © 1994 John Wiley & Sons, Inc.
Cultivation of an L‐lactate dehydrogenase mutant of Bacillus stearothermophilus in continuous culture with cell recycle
Authors: Martin R. San , Bushell D. , Leak D. J. , Hartley B. S. 
Published: December 2004

AbstractContinuous fermentation with cell recycle proved very effective in increasing the ethanol volumetric productivity of the thermophilic facultative anaerobe, Bacillus stearothermophilus strain LLD‐15, on sucrose at 70°C. When complete cell recycle was used, cell viability decreased after a few residence times and sucrose consumption was reduced. Operation using a constant bleed rate resulted in greater stability and higher ethanol volumetric productivities. A mathematical model based on maintenance energy requirements provided an adequate description of the system. © 1994 John Wiley & Sons, Inc.
Desulfurization of coal by microbial column flotation
Authors: Ohmura Naoya , Saiki Hiroshi 
Published: December 2004

AbstractTwenty‐three strains capable of oxidizing iron were isolated from coal and ore storage sites as well as coal and ore mines, volcanic areas, and hot spring. Four strains were found to have high iron‐oxidizing activity. One strain (T‐4) was selected for this experiment since the strain showed the fastest leaching rate of iron and sulfate from pyrite among the four strains. The T‐4 strain was assigned for Thiobacillus ferrooxidans from its cultural and morphological characteristics.Bacterial treatment was applied to column flotation. An increase of cell density in the microbial column flotation resulted in the increase of pyrite removal from a coal‐pyrite mixture (high sulfur imitated coal) with corresponding decrease of coal recovery. The addition of kerosene into the microbial column flotation increased the recovery of the imitated coal from 55% (without kerosene) to 81% (with 50 μL/L kerosene) with the reduction of pyrite sulfur content from 11% (feed coal) to 3.9% (product coal). The kerosene addition could reduce the pyritic sulfur content by collecting the coal in the recovery. However, the addition could not enhance separation of pyrite from the coal‐pyrite mixture, since pyrite rejection was not affected by the increase of the kerosene addition. An excellent separation was obtained by the microbial flotation using a long column which had a length‐diameter (L/D) ratio of 12.7. The long column flotation reduced the pyritic sulfur content from 11% (feed coal) to 1.8% (product coal) when 80% of the feed coal was recovered without the kerosene addition. The long column flotation not only attained an excellent separation but also reduced the amount of cells for desulfurization to as little as one‐tenth of the reported amount. © 1994 John Wiley & Sons, Inc.
On protein solubility in organic solvent
Authors: Chin Jennifer T. , Wheeler Sarah L. , Klibanov Alexander M. 
Published: December 2004

AbstractSolubility of a model protein, hen egg‐white lysozyme, was investigated in a wide range of neat nonaqueous solvents and binary mixtures thereof. All solvents that are protic, very hydrophilic, and polar readily dissolve more than 10 mg/mL of lysozyme (lyophilized from aqueous solution of pH 6.0). Only a marginal correlation was found between the lysozyme solubility in a non‐aqueous solvent and the letter's dielectric constant or Hildebrand solubility parameter, and no correlation was observed with the dipole moment. Lysozyme dissolved in dimethyl sulfoxide (DMSO) could be precipitated by adding protein nondissolving co‐solvents, although the enzyme had a tendency to form supersaturated solutions in such mixtures. The solubility of lysozyme, both in an individual solvent (1,5‐pentanediol) and in binary solvent mixtures (DMSO/acetonitrile), markedly increased when the pH of the enzyme aqueous solution prior to lyophilization was moved away from the proteins's isoelectric point. © 1994 John Wiley & Sons, Inc.