bluuebunny/crossref_metadata_2025_split huggingface dataset:
A Predictive Aggregate Transport Model for Microfiltration of Combined Macromolecular Solutions and Poly‐Disperse Suspensions: Testing Model with Transgenic Goat Milk
Authors: Baruah Gautam Lal , Couto Daniel , Belfort Georges 
Published: October 2003

AbstractTo meet the technical challenge of recovering human IgG fusion protein from transgenic whole goat milk at reasonable cost with high purity and yield, a predictive aggregate transport model for microfiltration has been developed (Baruah and Belfort, 2003). Here, to test the modelapos;s predictability of permeate flux and mass transport, a comprehensive series of experiments with varying wall shear rate, feed temperature, feed concentration, and module design are presented. A very good fit was obtained between the model predictions and measurements for a wide variety of experimental conditions. For microfiltration module design comparison, a linear hollow fiber module (representing current commercial technologies) gave lower permeation flux and higher yield than a helical hollow fiber module (representing the latest self‐cleaning methodology). These results are easily explained with the model that is now being used to define operating conditions for maximizing performance. The procedure described by the model is generalizable and can be used to obtain optimal filtration performance for applications other than milk.
Specific Oxygen Uptake Rate Variations during Batch Fermentation of Bacillus thuringiensis Subspecies kurstaki HD‐1
Authors: Rowe Gerald E. , Margaritis Argyrios , Wei Ning 
Published: October 2003

AbstractThe specific oxygen uptake rate (qO2, respiration rate) of Bacillus thuringiensis subsp. kurstaki HD‐1 was very high at inoculation and was found to decrease essentially monotonically throughout both vegetative growth phase and transition phase under different batch culture conditions. Average qO2 values decreased from 8–10 mmol/g h at 1 h after inoculation to less than 2 mmol/g h by the time growth ended. The results are shown to be consistent with the few previous reports on qO2 in B. thuringiensis in the literature but also novel in that this pattern of monotonic decline has not been described previously. Both pH control and EDTA in low concentration shortened the vegetative growth phase and reduced the 10 h biomass concentration. Using plots of qO2 versus specific growth rate, μ, biomass yield based on the oxygen used for growth, YmaxO2, was calculated for transition phase to be 0.041–0.047 g/mmol, consistent with literature values. The same plot also showed that the presence of EDTA resulted in an atypical qO2‐μ trajectory and apparently much higher biomass yield from the oxygen consumed.
Effect of Oxygen Delignification Operating Parameters on Downstream Enzymatic Hydrolysis of Softwood Substrates
Authors: Charles Nichola , Mansfield Shawn D. , Mirochnik Olga , Duff Sheldon J. B. 
Published: October 2003

AbstractEnhanced oxygen delignification of softwood pulp samples (taken upstream and downstream of a commercial oxygen delignification unit) improved the initial rate of enzymatic saccharification and overall yield of monomeric sugars by 62–82% and 76–80%, respectively. Laboratory‐scale experiments were used to examine the effect of a broad range of operating parameters (temperature, time, caustic concentration, and oxygen partial pressure) on the effectiveness of oxygen delignification. Using empirical models, kappa number (residual lignin content) was found to effectively predict final conversion to monomeric sugars. Application of oxygen delignification to sulfite mill knots resulted in smaller (20–25%) reduction in lignin content. However, using a combination of oxygen delignification and particle size reduction, up to 80% of the carbohydrate in the reject knots could be converted to fermentable sugars.
Perspectives for the Industrial Enzymatic Production of Glycosides
Authors: de Roode B. Mattheus , Franssen Maurice C. R. , Padt Albert van der , Boom Remko M. 
Published: October 2003

AbstractGlycosides are of commercial interest for industry in general and specifically for the pharmaceutical and food industry. Currently chemical preparation of glycosides will not meet EC food regulations, and therefore chemical preparation of glycosides is not applicable in the food industry. Thus, enzyme‐catalyzed reactions are a good alternative. However, until now the low yields obtained by enzymatic methods prevent the production of glycosides on a commercial scale. Therefore, high yields should be established by a combination of optimum reaction conditions and continuous removal of the product. Unfortunately, a bioreactor for the commercial scale production of glycosides is not available. The aim of this article is to discuss the literature with respect to enzymatic production of glycosides and the design of an industrially viable bioreactor system.
Supercritical Fluid Disruption of Ralstonia eutropha for Poly(β‐hydroxybutyrate) Recovery
Authors: Hejazi Parisa , Vasheghani‐Farahani Ebrahim , Yamini Yadollah 
Published: October 2003

AbstractA new method for disruption of Gram‐negative bacterium Ralstonia eutrophaby supercritical CO2 for poly(β‐hydroxybutyrate) (PHB) recovery is proposed. The effects of different parameters such as exposure time, pressure, temperature, volume of methanol as a modifier, and culture history on cell disruption efficiency were investigated using Taguchiapos;s statistical approach to determine optimum conditions. The optimum conditions for cell disruption and PHB recovery were as follows: exposure time, 100 min; pressure, 200 atm; temperature, 40 °C; volume of methanol, 0.2 mL. The cell culture time was less significant. At optimum conditions, the maximum efficiency of PHB recovery was found to be 89%. The proposed method is comparable with other recovery methods in terms of the percentage of PHB recovery, while it is environmentally more benign.
Micropatterned Substrates for the Growth of Functional Neuronal Networks of Defined Geometry
Authors: Vogt Angela K. , Lauer Lars , Knoll Wolfgang , Offenhäusser Andreas 
Published: October 2003

AbstractThe in vitro assembly of neuronal networks with control over cell position and connectivity is a fascinating approach not only for topics in basic neuroscience research but also in diverse applications such as biosensors and tissue engineering. We grew rat embryonic cortical neurons on patterned substrates created by microcontact printing. Polystyrene was used as a cell repellent background, onto which a grid pattern of physiological proteins was applied. We printed laminin and a mixture of extracellular matrix proteins and additionally both systems mixed with polylysine. Attachment of cells to the pattern with high fidelity as well as the formation of chemical synapses between neighboring cells on the pattern could be observed in all four cases, but cell attachment was strongly increased on samples containing polylysine. Neurons grown on patterned substrates had a membrane capacity smaller than that of neurons on homogeneously coated controls, which we attributed to the geometrical restrictions, but did not differ either in resting membrane potential or in the quality of synapses they formed. We therefore believe that the cells attach and differentiate normally on the pattern and form functional, mature synapses following the predefined geometry.
Effects of Three‐Dimensional Culturing on Osteosarcoma Cells Grown in a Fibrous Matrix: Analyses of Cell Morphology, Cell Cycle, and Apoptosis
Authors: Chen Chunnuan , Chen Kathryn , Yang Shang‐Tian 
Published: October 2003

AbstractOsteosarcoma cells were cultured in stirred tank bioreactors with either a fibrous matrix or nonporous microcarriers to study the environmental effects on cell growth, morphology, cell cycle, and apoptosis. Cell cycle and apoptosis were analyzed using flow cytometry and visualized using confocal laser scanning microscopy and fluorescence microscopy. The three‐dimensional (3‐D) fibrous culture had better cell growth and higher metabolic rates than the two‐dimensional (2‐D) microcarrier culture because cells in the fibrous matrix were protected from shear stress and had lower apoptosis and cell death even under suboptimal conditions (e.g., nutrient depletion). The polyester fibrous matrix used in this study also exhibited the capability of selectively retaining viable and nonapoptotic cells and disposing apoptotic and nonviable cells. Consequently, very few apoptotic cells were found in the fibrous matrix even in the long‐term (1 month) T‐flask culture. In the continuous culture with packed fibrous matrixes for cell support, most cells were arrested in the G1/G0 phase after 4 days. Decreasing the dissolved oxygen level from 60 to 10% air saturation did not significantly change cell cycle and apoptosis, which remained low at ∼15%. These results could explain why the fibrous bed bioreactor had good long‐term stability and was advantageous for production of non‐growth‐associated proteins by animal cell cultures.
Chimeric Gene Library Construction by a Simple and Highly Versatile Method Using Recombination‐Dependent Exponential Amplification
Authors: Ikeuchi Akinori , Kawarasaki Yasuaki , Shinbata Tomoya , Yamane Tsuneo 
Published: October 2003

AbstractA simple and efficient method for the construction of chimeric gene libraries termed RDA‐PCR (recombination‐dependent exponential amplification polymerase chain reaction) was developed by modifying polymerase chain reaction. A chimeric gene library is generated from homologous parental genes with additional primer‐annealing sequences at their “heads” and “tails”. Two primers (“skew primers”) are designed to exclusively anneal to either the heads of maternal genes or the tails of paternal genes. During the RDA‐PCR, short annealing/extension periods facilitate homologous recombination. The chimeric sequences can be exponentially amplified to form the chimeric gene library, whereas parental sequences without crossovers are not amplified. As a model, we constructed a chimeric gene library of yellow and green fluorescent protein (yfp and gfp, respectively). The crossover point profile of RDA‐PCR clones was compared with those obtained by (modified) family shuffling. PCR restriction fragment polymorphism (PCR‐RFLP) analysis of the RDA‐PCR clones showed a high content of chimeric genes in the library, whereas family shuffling required the modification using skew primers for selective enrichment of chimeric sequences. PCR‐RFLP analysis also indicated that the crossover points of RDA‐PCR chimeras were distributed over the entire protein‐coding region. Moreover, as few as 2 bp of the continual identity of nucleotides were found at the crossover points at high frequency (30% of the tested clones), suggesting that RDA‐PCR resulted in a higher diversity in crossover points than family shuffling.
Strategic Selection of Hyperthermophilic Esterases for Resolution of 2‐Arylpropionic Esters
Authors: Sehgal Amitabh C. , Kelly Robert M. 
Published: October 2003

AbstractHomologues to Carboxylesterase NP and Candida rugosa lipase, used for the chiral separation of racemic mixtures of 2‐arylpropionic methyl esters, were identified by BLAST searches of available genome sequences for hyperthermophilic microorganisms. Two potential candidates were identified: a putative lysophospholipase from Pyrococcus furiosus (Pfu‐LPL) and a carboxylesterase from Sulfolobus solfataricus P1 (Sso‐EST1). Although both enzymes showed hydrolytic preference toward the (S) methyl ester, only Sso‐EST1 yielded highly optically pure (S) naproxen (%eep ≥ 90) and was thus further investigated. Changes in pH or reaction time showed little improvement in %eep or E values with Sso‐EST1. However, the addition of 25% methanol resulted in a 25% increase in E. The effect of various cosolvents on the enantiomeric ratio showed no correlation with the log P or dielectric constant values of the solvent. However, an inverse relationship between E and the denaturation capacity (DC) of the water miscible cosolvents was observed. This was attributed to an increase in enzyme flexibility with increasing solvent DC values leading to a concomitant reduction in the resolving power of Sso‐EST1. The results here show that although bioinformatics tools can be used to select candidate biocatalysts for chiral resolution of 2‐arylpropionic esters, biochemical characterization is needed to definitively determine functional characteristics.
Production of Podophyllotoxin Using Cross‐Species Coculture of Linum flavum Hairy Roots and Podophyllum hexandrum Cell Suspensions
Authors: Lin Han‐wei , Kwok Kian H. , Doran Pauline M. 
Published: October 2003

AbstractNovel cross‐species coculture systems using Linum flavumhairy roots and Podophyllum hexandrum cell suspensions were applied for in vitro production of podophyllotoxin. The hairy roots and suspensions were cocultured in Linsmaier and Skoog medium in dual shake flasks and dual bioreactors. In separate experiments, coniferin feeding was shown to be an effective strategy for increasing the accumulation of podophyllotoxin in P. hexandrum suspensions. Because roots of L. flavum are a natural source of coniferin, hairy roots of this species were used in coculture with P. hexandrum to provide an in situ supply of coniferin. Compared with P. hexandrumsuspensions cultured alone in shake flasks or bioreactors, podophyllotoxin concentrations in cocultured P. hexandrum cells were increased by 240% and 72% in dual shake flask and dual bioreactor systems, respectively. The availability and stability of coniferin in the medium are the most likely factors limiting podophyllotoxin synthesis in coculture. Intensification of the coculture process is required to further improve total podophyllotoxin accumulation on a volumetric basis.
Predicted Unfolding Order of the 13 α‐Helices in the Catalytic Domain of Glucoamylase from Aspergillusawamori var. X100 by Molecular Dynamics Simulations
Authors: Liu Hsuan‐Liang , Wang Wen‐Chi 
Published: October 2003

AbstractThe unfolding mechanism of the 13 α‐helices in the catalytic domain of Aspergillus awamori var. X100 glucoamylase was investigated by 200 ps molecular dynamics simulations in explicit water with temperature jump technique. Rather than a simultaneous event, the unfolding of these 13 α‐helices followed a random ordered mechanism as α8→α1→α11→α7→α10→α3→α12→α13→α4→α5→α9→α6→α2. No significant relationships were found between the unfolding order and the length and the hydrophobicity of the helix. α‐Helix 8 located in the inner region of the catalytic domain was predicted to be the first helix to unfold, indicating that the destruction of the secondary structure motif was initiated from the inner region of the catalytic domain. The dynamic behavior of these α‐helices induced by increased kinetic energy during the unfolding process is considered to be similar to the expansion and compression of a series of springs under the influence of mechanical stress.
Tuning the Transcription Rate of Recombinant Protein in Strong Escherichiacoli Expression Systems through Repressor Titration
Authors: Striedner Gerald , Cserjan‐Puschmann Monika , Pötschacher Florentina , Bayer Karl 
Published: October 2003

AbstractThe main goal of this work was to develop a strategy that enables tuning of recombinant gene expression relative to the metabolic capacity of the host cell synthesis machinery. In the past, strong expression systems have been developed in order to maximize recombinant gene expression. However, these systems exert an extremely high metabolic burden onto the host cell, which may even lead to cell death. Hence, the period of recombinant gene expression is significantly reduced, and therefore, maximal yield cannot be attained. To extend the production phase and to achieve optimal yields, adjustment of recombinant gene expression by modulation of the transcription rate is required. To control transcription, we designed a feed regime, which continuously supplies limiting amounts of inducer in a constant ratio to biomass. For the accurate determination of appropriate amounts of inducer, a time shifted exponential substrate and inducer feed strategy has been developed. The potential of this metabolic and engineering integrated approach was proven in fed‐batch cultivation experiments using E. coliHMS174(DE3)(pET11ahSOD) as model system. Furthermore, our strategy enables the use of lactose as inducer, since its consumption can be compensated by appropriate feed profiles. The attained results fully comply with all requirements of industrial large scale cultivation and improve the applicability of strong expression systems.
Polymerization of Guaiacol and a Phenolic β‐O‐4‐Substructure by Trameteshirsuta Laccase in the Presence of ABTS
Authors: Rittstieg Klaus , Suurnäkki Anna , Suortti Tapani , Kruus Kristiina , Guebitz Georg M. , Buchert Johanna 
Published: October 2003

AbstractThe reaction of the monomeric lignin model compound guaiacol and the β‐O‐4‐type dimer erol (1–(4‐hydroxy‐3‐methoxyphenyl)–2(2‐methoxyphenoxy)‐propane‐1,3‐diol with laccase from Trametes hirsuta was studied in the presence of the mediator ABTS (2,2′‐azino‐di[3ethylbenzothiazoline‐6‐sulfonic acid]). The product mixtures were analyzed by means of aqueous‐phase size exclusion chromatography (SEC) with 50 mM NaOH as eluent. Interestingly, in the laccase‐catalyzed reaction with both substrates, the mediator not only functioned as an electron carrier but underwent coupling reactions with the substrate to give polymeric coupling products. The molecular weight of these copolymeric products was significantly higher than the molecular weight of products obtained without ABTS. After ultrafiltration, 33% and 21% of the initially applied ABTS could be found in the polymeric product fraction for the substrates guaiacol and erol, respectively, on the basis of nitrogen analysis. When ABTS was added to substrates after full laccase‐catalyzed polymerization, the reaction proceeded toward higher molecular weights.
Study of Process Variables in Industrial Acetic Fermentation by a Continuous Pilot Fermentor and Response Surfaces
Authors: Garrido‐Vidal Diego , Pizarro Consuelo , González‐Sáiz José M. 
Published: October 2003

AbstractThe modeling and optimization of industrial processes requires an intensive study of the factors involved. In this work, a continuous pilot system for studying the industrial process of acetic fermentation is developed. A Doehlert design is applied to the five variables involved in the pilot process. This experimental design allows reduction of the experimental burden and the maximum amount of information to be obtained, studying the factors at different levels depending on their significance. The experimental system provides a robust measure of the specific growth rate and the rates of substrates consumption and acetic acid production, related to the flow of effluent stream evaluated in the steady state. The results demonstrate the growth‐associated kinetics of substrates and product, and the yield factors are calculated with low values of variances for the coefficients, i.e., within the range 1–11%. The specific growth rate suits the quadratic model proposed. The response surfaces generated by the model are applied to explain the behavior of the bacterial growth and, therefore, the effects of the process variables studied over the acetic acid production. Very low levels of ethanol or oxygen make the acetification rate decrease, and a saturation effect with high levels of ethanol or oxygen is also deduced. The effects of the aeration rate, agitation, and overpressure suggest a kind of inhibition of the acetic acid production caused by the oxygen that has not been practically studied before. The temperature strengthens the inhibitory effect of the ethanol and the oxygen. The conclusions of this work consolidate the structure of a hybrid model for the acetic fermentation.
EASE Vectors for Rapid Stable Expression of Recombinant Antibodies
Authors: Aldrich Teri L. , Viaje Aurora , Morris Arvia E. 
Published: October 2003

AbstractOver the past 10 years, monoclonal antibodies and antibody fragments have become an increasingly important source of therapeutic molecules in the biotechnology industry. Drug development strategies rely on screening large numbers of candidate molecules in search of an optimized drug candidate. This strategy requires efficient production of ten to a few hundred milligrams of candidate molecules for screening in bioassays and animal models. Typically, this amount of recombinant protein expression involves large numbers of transient transfections or cloning of a recombinant cell line. Both of these approaches are time‐consuming and labor‐intensive. In this report, we describe the application of an EASE vector system that is capable of generating stable pools of transfected Chinese hamster ovary cells. These pooled populations of cells produce high quantities of antibody candidates without labor‐intensive cloning in a 3–5 week time frame. When an optimal drug candidate has been selected, pools generated with EASE‐containing vectors can also be used in subsequent cloning steps to make cell lines with improved expression levels. We demonstrate that EASE increases expression in nonamplified pools in addition to increasing amplification and viability of clonal cell lines generated with the EASE‐containing vectors compared with pools and cell lines generated without EASE.
Altered Glucose Transport and Shikimate Pathway Product Yields in E.coli
Authors: Yi Jian , Draths K. M. , Li Kai , Frost J. W. 
Published: October 2003

AbstractDifferent glucose transport systems are examined for their impact on phosphoenolpyruvate availability as reflected by the yields of 3‐dehydroshikimic acid and byproducts 3‐deoxy‐d‐arabino‐heptulosonic acid, 3‐dehydroquinic acid, and gallic acid synthesized by Escherichia coli from glucose. 3‐Dehydroshikimic acid is an advanced shikimate pathway intermediate in the syntheses of a spectrum of commodity, pseudocommodity, and fine chemicals. All constructs carried plasmid aroFFBR and tktA inserts encoding, respectively, a feedback‐insensitive isozyme of 3‐deoxy‐d‐arabino‐heptulosonic acid 7‐phosphate synthase and transketolase. Reliance on the native E. coli phosphoenolpyruvate:carbohydrate phosphotransferase system for glucose transport led in 48 h to the synthesis of 3‐dehydroshikimic acid (49 g/L) and shikimate pathway byproducts in a total yield of 33% (mol/mol). Use of heterologously expressed Zymomonas mobilis glf‐encoded glucose facilitator and glk‐encoded glucokinase resulted in the synthesis in 48 h of 3‐dehydroshikimic acid (60 g/L) and shikimate pathway byproducts in a total yield of 41% (mol/mol). Recruitment of native E. coli galP‐encoded galactose permease for glucose transport required 60 h to synthesize 3‐dehydroshikimic acid (60 g/L) and shikimate pathway byproducts in a total yield of 43% (mol/mol). Direct comparison of the impact of altered glucose transport on the yields of shikimate pathway products synthesized by E. coli has been previously hampered by different experimental designs and culturing conditions. In this study, the same product and byproduct mixture synthesized by E. coli constructs derived from the same progenitor strain is used to compare strategies for increasing phosphoenolpyruvate availability. Constructs are cultured under the same set of fermentor‐controlled conditions.