bluuebunny/crossref_metadata_2025_split huggingface dataset:
Inclined Sedimentation for Selective Retention of Viable Hybridomas in a Continuous Suspension Bioreactor
Authors: Batt Brian C. , Davis Robert H. , Kompala Dhinakar S. 
Published: March 2005

AbstractThe continuous separation of nonviable hybridoma cells from viable hybridoma cells by using a narrow rectangular channel that is inclined from the vertical has been investigated experimentally. The effectiveness of the settler in selectively retaining viable hybridomas in the bioreactor while permitting the removal of nonviable hybridomas has been shown to depend on the flow rate through the settler. Intermediate flow rates through the settler have been found to provide the highest removal of nonviable hybridomas relative to viable hybridoma retention. At high dilution rates through the chemostat, over 95% of the viable cells could be partitioned to the bottom of the settler while over 50% of the nonviable cells are removed through the top of the settler. This successful separation is due to the significantly larger size of the viable hybridomas than the nonviable ones. A continuous perfusion experiment was performed in which an external inclined settler was used to retain virtually all of the viable hybridomas in the culture, while selectively removing from the culture approximately 20% of the nonviable cells that entered the settler. A stable viable cell concentration of 1.0 × 107 cells/mL was achieved, as was an antibody productivity of over 50 μg/(mL‐day). These represent 3‐and 6‐fold increases, respectively, over the values obtained from a chemostat culture without cell retention.
Propagation of Recombinant Vaccinia Virus in HeLa Cells: Adsorption Kinetics and Replication in Batch Cultures
Authors: Chillakuru Rajeev A. , Ryu Dewey D. Y. , Yilma Tilahun 
Published: March 2005

AbstractThe influence of various culture parameters on infection and replication of recombinant vaccinia virus in HeLa cells was examined during various phases of viral replication. A modified form of the model of Valentine and Allison (Biochim. Biophys. Acta 1960, 40, 393–399) model was used to predict successfully the viral adsorption rates in cell suspensions. An experimentally determined aggregation factor, ϵ, was included in the model to account for deviations of the observed adsorption rates from those predicted by the earlier model. It was also shown that the ionic strength, ionic species, and serum proteins present in the medium significantly altered the adsorption kinetics of the virus. The lysosomotropic base chloroquine was found to enhance viral infection more than 2‐fold during the penetration step of viral infection. It was also demonstrated that cells infected during the exponential growth phase gave higher viral yields than those infected during the lag or stationary growth phases and the initial viral MOI did not significantly alter viral yields. Finally, it was demonstrated that viral infection of HeLa cells grown in 4‐L bioreactor batch cultures resulted in increased death and glucose uptake rates and significantly lower growth rates.
Reaction Rates for the Production of Selected Hormones by Solid‐Phase Synthesis
Authors: Wang Shen , Foutch Gary L. 
Published: March 2005

AbstractReaction rates have been measured for the production of selected bradykinins, angiotensins, and endorphins by solid‐phase synthesis. Reactor liquid concentrations are monitored by using a UV detector and flow cell. Experimentation has been limited to low excesses of the t‐Boc amino symmetrical anhydride. More than 500 attachments have been monitored. Most data obtained from resins with 1% cross‐linking show second‐order behavior with reaction rate constants between 0.5 and 8L/(mol‐s). The reaction rate is affected by chemical structures of both the attached amino symmetrical anhydride and the anchored amino terminus on the peptide fragment. Increasing reaction temperature and initial solution concentrations promotes reaction rate. The structure of the polymer support affects not only the reaction rate but also the observed reaction kinetics.
Solvent Selection Strategies for Extractive Biocatalysis
Authors: Bruce Lynda J. , Daugulis Andrew J. 
Published: March 2005

AbstractThis report follows the development of systematic solvent screening strategies for the identification of superior pure solvents and introduces techniques for the identification of effective coextractants. Specifically, methods to predict the biocompatibility and extractant capability of solvents are discussed. Biocompatibility is predicted by using heuristic data or the correlations between bioactivity and the logarithm of the partition coefficient of the solvent or the concentration of solvent in the cell membrane. A computer program, known as the extractant screening program or ESP, has been developed to effectively predict the behavior of virtually any product in any solvent/aqueous system. It is demonstrated that a biocompatible yet poor solvent can be mixed with a toxic solvent that has better extractant properties to yield a mixture with improved solvent characteristics that is still biocompatible. The fact that solvents do not mix in an ideal manner is exploited by using ESP to identify solvent mixtures that are still biocompatible at relatively high concentrations of toxic solvent.
Transport and Kinetics in Sandwiched Membrane Bioreactors
Authors: Jeong Yong S. , Vieth W. R. , Matsuura T. 
Published: March 2005

AbstractA bioreactor in which living yeast cells are sandwiched between an ultrafiltration membrane and a reverse osmosis membrane was constructed, and experiments were performed for the conversion of substrate glucose to product ethanol. A set of equations that include both transport through a series of barrier layers and bioreaction rate were developed to predict the performance of the sandwich bioreactor. The above equations were solved by using numerical values for the transport parameter and the bioreaction rate constant, and the results are compared with the experimental data.
Microscopic Visualization of Insect Cell‐Bubble Interactions. I: Rising Bubbles, Air‐Medium Interface, and the Foam Layer
Authors: Bavarian Farshad , Fan L. S. , Chalmers Jeffrey J. 
Published: March 2005

AbstractThrough the use of microscopic, high‐speed video technology, the interactions of two suspended insect cell lines, Trichoplusia ni (TN‐368) and Spodoptera frugiperda (SF‐9), with air and oxygen bubbles were studied. Events such as cell‐bubble attachment, cell‐bubble collision, cell transport into the foam layer, and trapping of cells in the foam layer are presented and discussed. Based on these observations and those in a companion paper (Chalmers, J. J.; Bavarian, F. Biotechnol. Prog. 1991, following paper in this issue) and the experimental and theoretical work of other researchers, several mechanisms of cell damage as a result of sparging are presented.
Microscopic Visualization of Insect Cell‐Bubble Interactions. II: The Bubble Film and Bubble Rupture
Authors: Chalmers Jeffrey J. , Bavarian Farshad 
Published: March 2005

AbstractIn this paper, the second in the series, the use of a microscopic, high‐speed video system to study the interactions of two suspended insect cells strains, Trichoplusia ni (TN‐368) and Spodoptera frugiperda (SF‐9), with rupturing bubbles is reported. Events such as the adsorption of cells onto the bubble film and the mechanism of bubble rupture were observed. On the basis of these observations and the experimental and theoretical work of other researchers on bubble rupture and cell death as a result of sparging, it is proposed that cells are killed by the rapid acceleration of the bubble film after rupture and the high levels of shear stress in the boundary layer flow associated with bubble jet formation.
Bromodeoxyuridine Labeling and Flow Cytometric Identification of Replicating Saccharomyces cerevisiae Cells: Lengths of Cell Cycle Phases and Population Variability at Specific Cell Cycle Positions
Authors: Dien Bruce S. , Srienc Friedrich 
Published: March 2005

AbstractAn immunofluorescent staining procedure has been developed to identify, with flow cytometry, replicating cells of Saccharomyces cerevisiae after incorporation of bromodeoxyuridine (BrdUrd) into the DNA. Incorporation of BrdUrd is made possible by using yeast strains with a cloned thymidine kinase gene from the herpes simplex virus. An exposure time of 4 min to BrdUrd results in detectable labeling of the DNA. The BrdUrd/DNA double staining procedure has been optimized and the flow cytometry measurements yield histograms comparable to data typically obtained for mammalian cells. On the basis of the accurate assessment of cell fractions in individual cell cycle phases of the asynchronously growing cell population, the average duration of the cell cycle phases has been evaluated. For a population doubling time of 100 min it was found that cells spend in average 41 min in the replicating phase and 24 min in the G2+M cell cycle period. Assuming that mother cells immediately reenter the S phase after cell division, daughter cells spend 65 min in the G1 cell cycle phase. Together with the single cell fluorescence parameters, the forward‐angle light scattering intensity (FALS) has been determined as an indicator of cell size. Comparing different temporal positions within the cell cycle, the determined FALS distributions show the lowest variability at the beginning of the S phase. The developed procedure in combination with multiparameter flow cytometry should be useful for studying the kinetics and regulation of the budding yeast cell cycle.
Regulation of Cellulase Synthesis in Acidothermus cellulolyticus
Authors: Shiang Ming , Linden James C. , Mohagheghi Ali , Grohmann Karel , Himmel Michael E. 
Published: March 2005

AbstractThe regulation of cellulase synthesis by induction and catabolite repression in the thermophilic, aerobic bacterium Acidothermus cellulolyticus was studied in batch fermentations. Various compounds, such as L‐sorbose, cyclic AMP (cAMP), L‐glucose, 2‐deoxyglucose (2‐DG), glucose 1‐phosphate (G‐1‐P), sophorose, salicin, sugar alcohols, and isopropyl thioglucoside (IPTGlu), were added along with Solka Floe to improve extracellular cellulase formation by the culture. When cAMP was added exogenously to A. cellulolyticus cultures in the concentration range of 0.01‐0.2 g/L, cAMP did not affect cell growth; however, cellulase yields were increased with increasing levels of cAMP. The enzyme production rates with the different levels of cAMP addition during Solka‐Floc fermentations were identical. L‐Sorbose, L‐glucose, 2‐DG, G‐1‐P, sophorose, IPTGlu, and sugar alcohols enhanced cellulase activity produced in the medium, but the starting time and the time required to reach the maximum enzyme activity were different in each condition. All these substances may function as moderators of cellulase synthesis. From the experimental results, only cellobiose, xylose, sophorose, and unknown soluble derivatives from cellulose were considered as inducers. In a possible regulatory mechanism of cellulase synthesis, the repressor, inducer, cAMP, and moderator may be all involved in controlling the rate and the yield of enzyme production.
Purification of Glycoproteins by Selective Transport Using Concanavalin‐Mediated Reverse Micellar Extraction
Authors: Paradkar Vikram M. , Dordick Jonathan S. 
Published: March 2005

AbstractA novel methodology for coupling liquid‐liquid extraction with affinity interaction has been developed to selectively and efficiently purify and separate glycoproteins. The basis for the separation is the selective extraction of glycoproteins from an aqueous solution into a reverse micellar organic phase by using concanavalin A (a sugar‐binding lectin) as a facilitative carrier. Specifically, horseradish peroxidase (a common glycoprotein) can be bound to concanavalin A in an aqueous phase and then extracted into an AOT‐isooctane organic phase with negligible loss in enzyme activity. Virtually no extraction of peroxidase occurs in the absence of concanavalin A. Electron spin resonance studies have shown that the large lectin‐glycoprotein complex (96 000 daltons) resides in a nonaqueous environment within the reverse micelle, perhaps at the surfactant, water‐pool interface; hence, extraction of the large complex is feasible. The facilitative extraction has been extended to selective transport of peroxidase from a mixture of peroxidase and alkaline phosphatase (a nonglycosylated protein). This results in an efficient separation strategy with a separation factor of 16.
Aminotransferase‐Catalyzed Conversion of D‐Amino Acids to L‐Amino Acids
Authors: Fotheringham Ian G. , Kidman Gene E. , Mcarthur Brian S. , Robinson Larry E. , Scollar Mark P. 
Published: March 2005

AbstractA microbial bioconversion process is described for the conversion of D‐amino acids or the corresponding racemic mixtures to L‐amino acids. The process comprises reacting a D‐amino acid or a D,L mixture of an amino acid with glucose and ammonia in the presence of an aminotransferase‐containing culture of Escherichia coli. The aminotransferase is only moderately stereoselective in converting D‐and L‐amino acids to the keto acids. The aminotransferase‐mediated conversion of α‐keto acid to L‐amino acid exhibits very high stereospecificity, thus rendering the conversion of D‐amino acids irreversible. Conversions of leucine, phenylalanine, and homophenylalanine to the corresponding L isomers are described.
Production of Fusion Protein SpA:: EcoRI in Batch Culture in a 60‐L Airlift Tower Loop Reactor
Authors: Brandes Lutz , Wu Xiaoan , Maschke Hans E. , Jürgens Heike , Reinhardt Birgit , Schügerl Karl 
Published: March 2005

AbstractEscherichia coli JM103 carrying the expression plasmid pMTC48, repressor plasmid pRK248, and protection plasmid pEcoR4 was grown in a 60‐L working volume airlift tower loop reactor on M9 minimal medium. Production of fusion protein SpA:: EcoRI was induced by a temperature shift from 30 to 38 (optimum), 40, or 42°C. The following parameters were monitored: cell mass concentration (X), total cell counts (TCC), number of colony‐forming units (CFU), concentrations of glucose, acetate, ethanol, pyruvate, lactate, succinate, amino acids, and ammonia, and soluble and total protein content, as well as product concentration (enzyme activity of the fusion protein), dissolved oxygen concentration, oxygen utilization rate (OUR), CO2 production rate (CPR), respiration quotient (RQ), and volumetric mass‐transfer coefficients (kLa). Product formation by temperature shift was only observed if LB concentrate was added to the culture at the same time the aeration rate was increased to avoid oxygen‐transfer limitation. No product accumulation was observed with glucose and ammonia supplementation. During gene expression, X and TCC increased, CFU decreased, acetate increased, and the primary metabolite (ethanol, pyruvate, lactate) concentrations as well as OUR and CPR passed a maximum while RQ changed only slightly. These facts indicate that, during gene expression, the metabolic activity of the cell passes a maximum, and after that it decreases. With increasing aeration rate, the volumetric productivity increased, but the specific productivity with respect to the cell concentration decreased.
Effect of Intraparticle Convection on the Chromatography of Biomacromolecules
Authors: Frey Douglas D. , Schweinheim Eberhard , Horváth Csaba 
Published: March 2005

AbstractThe effect of intraparticle convection in chromatographic columns packed with gigaporous particles (i. e., where dpore/dparticle > 10−2) on the band spreading of unretained biomacromolecules is investigated both experimentally and theoretically. A model is developed for the analysis of mass transfer in spherical particles of bidisperse pore structure when both convection and diffusion take place in the larger pores but only diffusion occurs in the smaller pores. The predictions of the model were experimentally verified. It is demonstrated that gigaporous particles have advantages over conventional porous particles (i. e., where dpore/dparticle < 10−3) for applications that do not require high resolving power, to bring about fast separation. This is because columns packed with gigaporous particles can be operated at high flow velocities without significant loss of efficiency due to the enhancement of mass transfer by intraparticle convection. The results of the model are used to examine the effectiveness of gigaporous column packings for rapid analytical chromatography and for the concentration and recovery of a dilute solute in a saturation‐regeneration cycle utilizing frontal chromatography.
Protein Purification with Novel Porous Sheets Containing Derivatized Cellulose
Authors: Manganaro James L. , Goldberg Bruce S. 
Published: March 2005

AbstractNovel porous sheets containing commercially available cellulosic ion exchange media of several different functionalities in a PVC matrix have been prepared and evaluated. The advantages of these materials are resistance to alkali, clean in place procedures and low nonspecific binding. These sheets thus provide well‐known and well‐characterized particulate chromatographic media in a porous sheet format. The porous sheet format permits use of short, squat geometries (stacked sheets) as well as tolerance of high pressure gradients. The net result is the ability to handle much higher flow rates (10‐fold or greater) than achievable by comparable cellulosic packed columns of the neat particulate media. Chromatographic separation of natural egg white proteins by a linear salt gradient on a single sheet of 1.3‐mm thickness was demonstrated. In addition, a very significant advantage of the porous sheet format is that it permits the fabrication of devices which are not possible with particulate media. An example of such a device is the Acti‐Mod Spiral Module. In the spiral module a spiral flow channel is formed by wrapping the porous sheet material around a mandrel. Embossed ribbing in the porous sheet provides channel spacing. The small open channel accommodates high flow rates of biological suspensions while the porous sheet walls selectively remove components of the suspension.
Growth of Plant Root Cultures in Liquid‐ and Gas‐Dispersed Reactor Environments
Authors: Mckelvey Scott A. , Gehrig Jason A. , Hollar Kathryn A. , Curtis Wayne R. 
Published: March 2005

AbstractThe growth of Agrobacterium transformed “hairy root” cultures of Hyoscyamus muticus was examined in various liquid‐ and gas‐dispersed bioreactor configurations. Reactor runs were replicated to provide statistical comparisons of nutrient availability on culture performance. Accumulated tissue mass in submerged air‐sparged reactors was 31% of gyratory shake‐flask controls. Experiments demonstrate that poor performance of sparged reactors is not due to bubble shear damage, carbon dioxide stripping, settling, or flotation of roots. Impaired oxygen transfer due to channeling and stagnation of the liquid phase are the apparent causes of poor growth. Roots grown on a medium‐perfused inclined plane grew at 48% of gyratory controls. This demonstrates the ability of cultures to partially compensate for poor liquid distribution through vascular transport of nutrients. A reactor configuration in which the medium is sprayed over the roots and permitted to drain down through the root tissue was able to provide growth rates which are statistically indistinguishable (95% T‐test) from gyratory shake‐flask controls. In this type of spray/trickle‐bed configuration, it is shown that distribution of the roots becomes a key factor in controlling the rate of growth. Implications of these results regarding design and scale‐up of bioreactors to produce fine chemicals from root cultures are discussed.
Measurement of Diffusion Coefficients in Gels Using Holographic Laser Interferometry
Authors: Gustafsson Nils Ove , Westrin Bengt , Axelsson Anders , Zacchi Guido 
Published: March 2005

AbstractThe accuracy and precision of holographic interferometry as a method to measure diffusion coefficients in gels are investigated both experimentally and theoretically. The standard deviations in the experimentally determined diffusion coefficient for ethanol in 4% (w/v) agarose gel were 3.3% for diffusion into the gel and 6.1% for diffusion out of the gel. These are in good agreement with the standard deviations obtained using Monte Carlo simulations. Systematic errors derived from an assumption of constant diffusion coefficients were also investigated.
Surface Area and Anchorage‐Dependent Growth of Chinese Hamster Ovary Cells
Authors: Ruaan R.‐C. , Tsai G.‐J. , Tsao G. T. 
Published: March 2005

AbstractCell growth can be affected by many factors. For study of the cell responses under various environmental changes, the traditional method using batch cultures may not provide precise answers. It sometimes even leads to misapprehension. A flow system was therefore designed to eliminate the variations in chemical environment during the cell growth. An anchorage Chinese hamster ovary (CHO) cell line was chosen for study under this system. With the help of time‐lapse video recording and image analysis, it was found that the exposed surface area (Ae) of Chinese hamster ovary (CHO) cells was significantly affected by the serum concentration (S) in the medium. Cells appeared to be larger when cultivated in medium of low serum content. Comparing the growth rates of cells on different substrata, we have found that, although they were different at a given serum concentration, the growth curves could be correlated to a Monod type equation by replacing the substrate term with AeS. Furthermore, when we studied the effect of density‐dependent inhibition of cell growth, we found that there exists a minimum value of AeS required for stimulating cell proliferation. Once AeS falls below this value, cell proliferation stops. These observations seemed to indicate that the interaction between serum factors and cell surface, i.e., AeS, is an important parameter of the growth of CHO cells.
Kinetics of Hepatocyte Spheroid Formation
Authors: Peshwa Madhusudan V. , Wu Florence J. , Follstad Brian D. , Cerra Frank B. , Hu Wei‐Shou 
Published: March 2005

AbstractCultured hepatocytes have been explored for use in a bioartificial liver. Spheroids formed by cultured hepatocytes exhibit enhanced liver‐specific functions. The kinetics of spheroid formation, using rat hepatocytes, was studied on positively charged surfaces that were either uncoated or coated with collagen or (hydroxyethyl)methacrylate (HEMA). Optimal spheroid formation was obtained on positively charged (Primaria) surfaces at inoculum densities in the range of (3–9) × 104 cells/cm2. Cells initially attached and spread out on the surface. Subsequent retraction led to the emergence of small clumps of cells attached to the surface, from which spheroids formed and shed off into suspension. The process of spheroid formation took more than 72 h and was accompanied by a decrease in the surface area occupied by attached cells. Optical sectioning of fluorescently stained spheroids using confocal microscopy indicated that most of the cells in the spheroid were viable. Spheroids also maintained a constant albumin synthesis rate for over 7 days in culture. Spheroid formation was evaluated in terms of the changes in spheroid diameter, the surface area covered by attached cells, and the total protein content of the fraction of cells that formed spheroids. The quantitative methodologies developed were used to assess the effect of inoculum cell concentration on spheroid formation and to evaluate the kinetics of spheroid formation on different surfaces both favorable and nonfavorable to spheroid formation.