bluuebunny/crossref_metadata_2025_split huggingface dataset:
Enhancement of monoclonal antibody yield by hybridoma fed‐batch culture, resulting in extended maintenance of viable cell population
Authors: Bushell M. E. , Bell S. L. , Scott M. F. , Spier R. E. , Wardell J. N. , Sanders P. G. 
Published: December 2004

AbstractHybridoma batch cultures were extended using feed formulations based on nutrient consumption measured during different batch culture phases when (a) growth but negligible antibody production was taking place; (b) maximum antibody production rate and declining viable cell growth rate were observed. Strategy (a) was the more successful (2.8‐fold compared with 1.8‐fold antibody titer increase) and maintained cell viability for longer. Analysis of the effects of omitting individual amino acids yielded results which were consistent with those from the feeding experiment © 1994 John Wiley & Sons, Inc.
A mathematical model for the leukocyte filtration process
Authors: Bruil Anton , Beugeling Tom , Feijen Jan 
Published: December 2004

AbstractLeukocyte filters are applied clinically to remove leukocytes from blood. In order to optimize leukocyte filters, a mathematical model to describe the leukocyte filtration process was developed by modification of a general theoretical model for depth filtration. The model presented here can be used to predict the time‐dependent leukocyte filtration as a function of cell–cell interaction in the filter, filter efficiency, filter capacity, filter dimensions, and leukocyte concentration in the suspension applied to the filter. The results of different leukocyte filtration experiments previously reported in the literature could be well described by the present model. © 1995 John Wiley & Sons, Inc.
Metal recognition by in‐vitro selection
Authors: Gupta Kanika , Yin John 
Published: December 2004

AbstractWe propose an in vitro selection strategy to identify bacteriophage variants that recognize metal ions in solution. In 6 M urea, phage T7 loses 99.9% of its activity in less than 5 min. Inactivation is accelerated by gold, but slowed by zinc and magnesium. Selection of phage over five generations in the presence of gold, zinc, and magnesium increases phage half‐lives 4‐, 10‐, and 70‐fold, respectively. As selections are repeated, phage become increasingly dependent on the specific metal employed in the selection, indicating the suitability of the strategy for optimization of metal‐ion recognition. © 1995 John Wiley & Sons, Inc.
Determination of the respiration quotient in mammalian cell culture in bicarbonate buffered media
Authors: Bonarius Hendrik P. J. , de Gooijer Cornelis D. , Tramper Johannes , Schmid Georg 
Published: December 2004

AbstractThe 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.
Effects of substrate branching characteristics on kinetics of enzymatic depolymerization of mixed linear and branched polysaccharides: II. Amylose/glycogen α‐amylolysis
Authors: Park Jun T. , Rollings James E. 
Published: December 2004

AbstractEnzymatic depolymerization of polysaccharides with α‐amylase has been studied in mixed aqueous dimethylsulfoxide (DMSO)/water solvents. Polysaccharide substrate chemical compositions, configurational structures, and bonding pattersn are known to affect observed enzymatic reaction kinetics. The branching structures of polysaccharides and their effects on the kinetic mechanisms of depolymerization reactions via endo‐acting hydrolyzing enzyme was studied via size exclusion chromatography coupled to low angle laser light scattering (SEC/LALLS). The glycogen branching structure is a heterogeneously distributed “cluster” structure rather than a homogeneously distributed “treelike” structure. The action pattern of α‐amylase on glycogen, which is composed of highly branched clusters, as end‐products, has a “pseudo‐exo‐attack” in contrast to an expected “endoattack” as seen in the hydrolysis of amylose or amylopectin substrates. These effects of branched substrates for mixed amylose/glycogen α‐amylolysis have been predicted and demonstrated by both experimental and theoretical analysis using the kinetic model presented in this report. The “lumped” kinetic model employed, assumes that the enzyme simultaneously attacks both linear and branched substrates. In general, excellent agreement between the model predictions and the experimental observations, both qualitatively and quantitatively, was obtained. © 1995 John Wiley & Sons, Inc.
Protein extration from an aqueous phase into a reversed micellar phase: Effect of water content and reversed micellar composition
Authors: Hilhorst R. , Sergeeva M. , Heering D. , Rietveld P. , Fijneman P. , Wolbert R. B. G. , Dekker M. , Bijsterbosch B. H. 
Published: December 2004

AbstractIn the system composed of the cationic surfactant TOMAC (10 mM), the nonionic (co)surfactant Rewopal HV5 (2 mM), and octanol (0.1% v/v) in isooctane, reversed micelles are formed upon contact with an aqueous phase containing 50 mM ethylene diamine. α‐Amylase can be transferred from the aqueous phase into reversed micelles in the pH range 9.5 to 10.5 and re‐extracted into a second aqueous phase of different composition. The size of the reversed micelles (as reflected in the water content of the organic phase) can be varied by changes in percentage of octanol, type of counterion in the aqueous phase, or in the number of ethoxylate head groups of the nonionic surfactant. An increase in size results in transfer at lower pH values. Experiments in which the charge density in the reversed micellar interface was changed by incorporation of charged derivatives of the nonionic surfactant, without influencing the water content, revealed that an increased charge density facilitated transfer, resulting in a broader transfer profile. Replacement of TOMAC by other quaternary ammonium surfactants differing in number and length of tails revealed that, of the 14 surfactants tested, only 2 gave appreciable amounts of transfer. The amount of transfer is related to the dynamics of phase separation of the surfactants: those giving a poor phase separation inactivate the enzyme. This inactivation is caused by electrostatic interactions between the charged surfactant head groups and charged groups on the enzyme. Electrostatic interactions are the first step of transfer, and can result in either incorporation in a reversed micelle, or, if reversed micelle formation is slow, in enzyme inactivation. © 1995 John Wiley & Sons, Inc.
The viability of Scenedesmus bicellularis cells immobilized on alginate screens following nutrient starvation in air at 100% relative humidity
Authors: Kaya Valentino M. , Picard Gaston 
Published: December 2004

AbstractThe 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.
Effect of single nutrient limitation of poly‐β‐hydroxybutyrate molecular weight distribution in alcaligens europhus
Authors: Asenjo J. A. , Schmidt A. S. , Andersen P. R. , Andrews B. A. 
Published: December 2004

AbstractThe effect of magnesium and phosphate limitation on the molecular weight distribution of poly‐β‐hydroxybutyrate (PHB) in Alcaligens europhus in cotinuons culture has been stuied. Conditions of nitrogen limitation both with glucose excess (above ca. 20 g/L) and without excess were investigated Under N‐limitation and glucose excess, Mw decreases when the magnesium content is decreased below 50% (19.7 mg/L) of the basal medium content; this also results in a broadenng of molecular weight distribution (Mw/Mn) from 2 to 5 and a decrease in Mw fron 2 × 106 to 0.9 × 106. Below 20% of the basal content of magnesium (7.9 mg/L) these two trends were reversed. This behaviour was not observed in the absence of glucose excess, phshate had virtually no effect on PHB Mw or its distribution, whereas wih no (or little) glucose excess Mw of the PHB decreased with phosphate concentrations below 50% of the basal level (0.705 g/L). Hence, in continuous or fed‐batch cultures, in addition to nitrogen limitation to alklow for PHB accumulation, it is necesary to control both the addition of glucose (no excess) and also to maintain magnesium limitation (ca. 25% of basal medium level, 9.9 mg/L) and phosphate above 50% of he basal level (0.705 g/L). Thus, when broadening of molecular weight destribution (increase in Mw/Mn) is observed at the end of fed‐batch culture it is probably caused by phosphate limitation and/or glucose excess. © 1995 John Wiley & Sons, Inc.
Effect of different cell culture conditions on the polypeptide integrity and N‐glycosylation of a recombinant model glycoprotein
Authors: Gawlitzek Martin , Conradt Harald S. , Wagner Roland 
Published: December 2004

AbstractThe effect of different short‐term controlled cell culture conditions on the product quality of a genetically engineered human interleukin‐2 N‐glycosylation variant protein expressed from a baby hamster kidney cell line (BHK‐21) has been investigated. A perfused 2‐L stirred tank reactor was used. Products purified from the culture supernatant of cells grown under experimentally initiated nutrient limitations (glucose, amino acids, pO2) were characterized by their HPLC‐elution profile, SDS‐PAGE and western blotting, amino acid sequencing as well as for their N‐linked carbohydrates, using “HPAEC‐PAD fingerprinting” and methylation analysis. The glycoprotein products secreted from cells under the different culture conditions (kept for 24 h, after an adaption time period of 48 h) showed an almost identical oligosaccharide pattern. By contrast, short‐term changes of the culture condition led to considerable differences in the ratio of glycosylated to unglycosylated protein forms. Significant amounts of NH2‐terminally truncated polypeptide forms were observed. They lacked proponderantly the first two amino acids; however, under certain culture conditions forms lacking up to eight NH2‐terminal amino acids were detected. © 1995 John Wiley & Sons, Inc.
The influence of electrostatic interactions on partition in aqueous polyethylene glycol/dextran biphasic systems: Part II
Authors: Schluck A. , Maurer G. , Kula M.‐R. 
Published: December 2004

AbstractIn aqueous polyethylene glycol/dextran two‐phase systems, the hydrophobicity, free volume, surface tension, and interfacial tension of the phases in equilibrium were measured as a function of pH and ionic strength. These parameters were found to change with pH, but the pattern and magnitude cannot explain the unusual partition of charged macromolecules, observed previously. The electrostatic potential difference was determined by a new experimental approach based on the measurement of the pH difference between the phases at equilibrium. In polyethylene glycol/dextran systems containing sodium chloride as ionized species, the electrostatic potential is not constant in the pH range 2 to 11. The partition behavior of charged macromolecules and its dependence on pH can be explained by the combined action of charge and phase potential. This conclusion was tested with poly‐L‐glutamate, which partitioned as predicted and in a pattern opposite to positively charged macro‐ molecules. © 1995 John Wiley & Sons, Inc.
Improvement of Escherichia coli microaerobic oxygen metabolism by Vitreoscilla hemoglobin: New insights from NAD(P)H fluorescence and culture redox potential
Authors: Tsai Philip S. , Rao Govind , Bailey James E. 
Published: December 2004

AbstractOn‐line NAD(P)H fluorescence and culture redox potential (CRP) measurements were utilized to investigate the role of Vitreoscilla hemoglobin (VHb) in perturbing oxygen metabolism of microaerobic Escherichia coli Batch cultures of a VHb‐synthesizing E. coli strain and the iso‐genic control under fully aerated conditions were subject to several high/low oxygen transitions, and the NAD(P)H fluorescence and CRP were monitored during these passages. The presence of VHb decreased the rate of net NAD(P)H generation by 2.4‐fold under diminishing oxygen tension. In the absence of aeration, the strain producing VHb maintained a steady NAD(P)H level 1.8‐fold less than that of the control, indicating that the presence of VHb keeps E. coli in a more oxidized state under oxygen‐limited conditions. Estimated from CRP, the oxygen uptake rates near anoxia were 25% higher for cells with VHb than those without. These results suggest that VHb‐expressing cells have a higher microaerobic electron transport chain turnover rate. To examine how NAD(P)H utilization of VHb‐expressing cells responds to rapidly changing oxygen tension, which is common in large‐scale fermentations, we pulsed air intermittently into a cell suspension and recorded the fluorescence response to the imposed dissolved oxygen (DO) fluctuation. Relative to the control, cells containing VHb had a sluggish fluorescence response to sudden changes of oxygen tension, suggesting that VHb buffers intracellular redox perturbations caused by extracellular DO fluctuations.© John Wiley & Sons, Inc.
Continuous monitoring of nitrogenase activity in Azotobacter vinelandii fermentation using off‐gas mass spectrometry
Authors: Prior John J. , Christie Patricia D. , Murray Robert J. , Orme‐Johnson William H. , Cooney Charles L. 
Published: December 2004

AbstractThis study evaluated the feasibility of monitoring nitro‐genase activity in situ through measurement of N2 uptake rate (NUR) using off‐gas mass spectrometry. Four 50‐L cultures of Azotobacter vinelandii were grown aer‐obically in nitrogen‐free medium to cell densities of 1.0‐1.3gL−1 magnetic‐sector mass spectrometer was used to monitor NUR along with other gas exchange rates. The small specific uptake rate (1.2 mmol g−1 h−1) and low cell density were found to lead to a NUR below the measurement accuracy limits under normal conditions. An operating strategy and feed gas mixture (40% O2, 45% N2 15% Ar) were designed to improve the signal‐to‐noise ratio while maintaining dissolved O2 and N2 levels in desired ranges. The fraction of N2 removed from the air stream was increased approximately 5‐fold from 0.2% to 1.0% and the measurement noise was reduced 25‐fold from a baseline of ±5to ±0.2 mmol L−1 h−1. The NUR measurements were compared against in vivo and in vitro acetylene reduction assays as well as on‐line cell growth rate measurements. While electron transfer requirements predict an NUR‐to‐acetylene reduction rate ratio of 0.33, measured ratios for the in vivo and in vitro assays were 0.8 and 0.44, respectively. This suggests that other rate‐limiting steps were present in the case of the in vivo assay. In accordance with reports in the literature, no concomitant hydrogen evolution was detected. This is the first reported continuous and direct measurement of NUR in fermentation and demonstrates a novel approach for improving measurement accuracy through rational adjustment of operating conditions. The technique has potential to provide useful insight for development and control of microbial nitrogen fixation processes.© John Wiley & Sons, Inc.
Protein‐free human–human hybridoma cultures in an intercalated‐spiral alternate‐dead‐ended hollow fiber bioreactor
Authors: Brotherton John D. , Chau Pao C. 
Published: December 2004

AbstractBatch 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.
Sustained and constitutive high levels of protein production in continuous cultures of bacillus subtilis
Authors: Vierheller Chenzhao , Goel Akshay , Peterson Michael , Domach Michael M. , Ataai Mohammad M. 
Published: December 2004

AbstractThe 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.
Domain E of Bacillus macerans cyclodextrin glucanotransferase: An independent starch‐binding domain
Authors: Dalmia Bipin K. , Schütte Kai , Nikolov Zivko L. 
Published: December 2004

AbstractThe starch‐binding domains of glucoamylase I (SBD of GA‐I) from Aspergillus awamori and of cyclodextrin glucanotransferase (domain E of CGTase) from Bacillus macerans were fused to the C‐terminus of β‐galactosidase (β‐gal) The majority of the fusion proteins produced in Escherichia coli were found as inclusion bodies. Active fusion proteins were purified by partial solubilization of the inclusion bodies with 2 M urea followed by affinity chromatography. Adsorption isotherms of purified fusion proteins on corn starch and cross‐linked amylose were generated. The β‐gal fusion proteins had similar affinities for cross‐linked amylose and corn starch but significantly different saturation capacities on corn starch. The adsorption and elution data from the potato starch column as well as the adsorption isotherms of p‐gal‐domain E fusion protein (BDE109) on corn starch and cross‐linked amylose demonstrated that domain E of CGTase is an independent domain, which retained its starch‐binding activity when separated from the other four (A–D) domains in CGTase. © 1995 John Wiley & Sons Inc.
Characterization of Penicillium chrysogenum physiology in submerged cultures by color and monochrome image analysis
Authors: Vanhoutte B. , Pons M. N. , Thomas C. R. , Louvel L. , Vivier H. 
Published: December 2004

AbstractAlthough filamentous microorganisms are widely used in industrial fermentation processes, their growth and differentation are not yet fully understood, because their biomass is structured, and therefore difficult to descrbie and to quantify. This lack of appropriate tools can hinder the optimization and control of the fermentation. A quantitative image analysis method was therefore developed for characterizing the physiology of the penicillin‐producing mold Penicillium chrysogenum. This method is based on a differntial staining procedure showing six physiological states: growing material, three differentiated states characterzied by an increasing granulation, a highly vacuolized state, and dead segments having lost their cytoplasm. The image analysis software, with versions written for monochrome and color images, consisted of a semiautomatic binary mask computation step and a fully automatic segmentation step based on a fuzzy classification. © 1995 John Wiley & Sons, Inc.