bluuebunny/crossref_metadata_2025_split huggingface dataset:
Computational Fluid Dynamics Modeling of Steady‐State Momentum and Mass Transport in a Bioreactor for Cartilage Tissue Engineering
Authors: Williams Kenneth A. , Saini Sunil , Wick Timothy M. 
Published: October 2002

AbstractComputational fluid dynamics (CFD) models to quantify momentum and mass transport under conditions of tissue growth will aid bioreactor design for development of tissue‐engineered cartilage constructs. Fluent CFD models are used to calculate flow fields, shear stresses, and oxygen profiles around nonporous constructs simulating cartilage development in our concentric cylinder bioreactor. The shear stress distribution ranges from 1.5 to 12 dyn/cm2 across the construct surfaces exposed to fluid flow and varies little with the relative number or placement of constructs in the bioreactor. Approximately 80% of the construct surface exposed to flow experiences shear stresses between 1.5 and 4 dyn/cm2, validating the assumption that the concentric cylinder bioreactor provides a relatively homogeneous hydrodynamic environment for construct growth. Species mass transport modeling for oxygen demonstrates that fluid‐phase oxygen transport to constructs is uniform. Some O2 depletion near the down stream edge of constructs is noted with minimum pO2 values near the constructs of 35 mmHg (23% O2 saturation). These values are above oxygen concentrations in cartilage in vivo, suggesting that bioreactor oxygen concentrations likely do not affect chondrocyte growth. Scale‐up studies demonstrate the utility and flexibility of CFD models to design and characterize bioreactors for growth of tissue‐engineered cartilage.
Reactor Operation and Scale‐Up of Whole Cell Baeyer‐Villiger Catalyzed Lactone Synthesis
Authors: Doig Steven D. , Avenell Philip J. , Bird Paul A. , Gallati Patrick , Lander Katie S. , Lye Gary J. , Wohlgemuth Roland , Woodley John M. 
Published: October 2002

AbstractThe recombinant whole cell biocatalyst Escherichia coli TOP10 [pQR239], expressing cyclohexanone monooxygenase from Acinetobacter calcoaceticus NCIMB 9871, was used in 1.5‐ and 55‐L fed‐batch processes to oxidize bicyclo[3.2.0]hept‐2‐en‐6‐one to its corresponding regioisomeric lactones, (–)‐(1 S,5 R)‐2‐oxabicyclo[3.3.0]oct‐6‐en‐3‐one and (–)‐(1 R,5 S)‐3‐oxabicyclo[3.3.0]oct‐6‐en‐2‐one. By employing a bicyclo[3.2.0]hept‐2‐en‐6‐one feed rate below that of the theoretical volumetric biocatalyst activity (275 μmol·min−1·L−1), the reactant concentration in the bioreactor was successfully maintained below the inhibitory concentration of 0.2–0.4 g·L−1. In this way approximately 3.5 g·L−1 of the combined regioisomeric lactones was produced with a yield of product on reactant of 85–90%. The key limitation to the process was shown to be product inhibition. This process was scaled up to 55 L, producing over 200 g of combined lactone product. Using a simple downstream process (centrifugation, adsorption to activated charcoal, 5‐fold concentration with ethyl acetate elution, and silica gel chromatography), we have shown that the two regioisomeric lactone products could be isolated and purified at this scale.
Multiple Substrate Growth Kinetics of Leptothrix discophoraSP‐6
Authors: Yurt Nurdan , Sears John , Lewandowski Zbigniew 
Published: October 2002

AbstractThe growth parameters of Leptothrix discophora SP‐6 were quantified on the basis of the steady‐state concentrations and utilization rates of pyruvate, dissolved oxygen, and concentration of microorganisms in a chemostat operated at 25 °C, pH 7.2, and an agitation rate of 350 rpm. The results showed that the microbial growth was limited by both pyruvate and dissolved oxygen. A combined growth kinetics model using Monod growth kinetics for pyruvate and Tessier growth kinetics for oxygen showed the best correlation with the experimental data when analyzed using an interactive multiple substrate model. The growth kinetics parameters and the respective confidence limits, estimated using the Monte Carlo simulation, were μmax = 0.576 ± 0.021 h−1, KsMp = 38.81 ± 4.24 mg L−1, KsTo = 0.39 ± 0.04 mg L−1, YX/p = 0.150 (mg microorganism mg−1 pyruvate), YX/o = 1.24 (mg microorganism mg−1 oxygen), the maintenance factors for pyruvate and oxygen were mp = 0.129 (mg pyruvate consumed mg−1 microorganism h−1) and mo = 0.076 (mg oxygen consumed mg−1 microorganism h−1), respectively.
Decreased Protein Expression and Intermittent Recoveries in BiP Levels Result from Cellular Stress during Heterologous Protein Expression in Saccharomyces cerevisiae
Authors: Kauffman Kenneth J. , Pridgen Eric M. , Doyle Francis J. , Dhurjati Prasad S. , Robinson Anne Skaja 
Published: October 2002

AbstractCells are inherently robust to environmental perturbations and have evolved to recover readily from short‐term exposure to heat, pH changes, and nutrient deprivation during times of stress. The stress of unfolded protein accumulation has been implicated previously in low protein yields during heterologous protein expression. Here we describe the dynamics of the response to this stress, termed the unfolded protein response (UPR), during the expression of the single chain antibody 4–4–20 (scFv) in Saccharomyces cerevisiae. Expression of scFv decreased the growth rate of yeast cells whether the scFv was expressed from single‐copy plasmids or integrated into the chromosome. However, the growth rates recovered at longer expression times, and surprisingly, the recovery occurred more quickly in the high‐copy integration strains. The presence of a functional UPR pathway was necessary for a recovery of normal growth rates. During the growth inhibition, the UPR pathway appeared to be activated, resulting in decreased intracellular scFv levels and intermittent recovery of the chaperone BiP within the endoplasmic reticulum. Intracellular scFv was observed primarily in the endoplasmic reticulum, consistent with activation of the UPR pathway. Although the intracellular scFv levels dropped over the course of the expression, this was not a result of scFv secretion. A functional UPR pathway was necessary for the drop in intracellular scFv, suggesting that the decrease was a direct response of UPR activation. Taken together, these results suggest that control of heterologous gene expression to avoid UPR activation will result in higher production levels.
Protein Purification via Aqueous Two‐Phase Extraction (ATPE) and Immobilized Metal Affinity Chromatography. Effectiveness of Salt Addition To Enhance Selectivity and Yield of GFPuv
Authors: Li Yi , Beitle Robert R. 
Published: October 2002

AbstractThis study illustrates the compatibility and complementary nature of aqueous two‐phase extraction (ATPE) and immobilized metal affinity chromatography (IMAC) in a general recovery scheme. The purification of green fluorescent protein (GFPuv) from extracts of Eschericia coli was investigated using a combination of these two techniques. High molarity of sodium chloride was found effective in increasing selectivity, with the promotion of hydrophobic interaction the probable mechanism that drove the target protein to a particular phase in ATPE, as well as that which enhanced GFPuv adsorption in IMAC. Moreover, the similar salt condition allows the direct application of the GFPuv‐containing phase to the IMAC column without additional adjustment step. A simple screen of conditions was therefore performed to generate a favorable two‐step purification scheme for GFP leading to an overall high purity.
Isoprene Formation in Bacillus subtilis: A Barometer of Central Carbon Assimilation in a Bioreactor?
Authors: Shirk Megan C. , Wagner William P. , Fall Ray 
Published: October 2002

AbstractIsoprene (2‐methyl‐1,3‐butadiene) is a volatile hydrocarbon of uncertain function in Bacillus subtilis, and we hypothesized that it is an overflow metabolite produced during excess carbon utilization. Here we tested this idea for phase 2 of isoprene release, a phase that occurs during extracellular acetoin accumulation and its reassimilation. Phase 2 isoprene formation could be disrupted in three different ways, all related to acetoin metabolism. Disruption of a gene essential for acetoin biosynthesis (acetolactic acid synthase, alsS) blocked acetoin formation and led to cessation of phase 2 isoprene formation as well as a variety of pleiotropic effects related to loss of pH control. Growth of the alsS mutant with external pH control reversed most of these effects. Disruption of acetoin catabolism (acetoin dehydrogenase, acoA), also eliminated phase 2 isoprene formation and caused cells to transition directly from phase 1 to phase 3; the latter is attributed to amino acid catabolism. A third alteration of acetoin metabolism was detected in the widely used strain 168 ( trpC2) but not in strain MS175, a trpC mutant constructed in the Marburg strain genetic background. Strain 168 exhibited slow acetoin assimilation compared to that of MS175 or the parental strain, with little or no isoprene formation during this growth phase. These findings support the idea that isoprene release occurs primarily when the rate of carbon catabolism exceeds anabolism and that this volatile hydrocarbon is a product of overflow metabolism when precursors are not required for higher isoprenoid biosynthesis. It is suggested that isoprene release might serve as a useful barometer of the rise and fall of central carbon fluxes during the growth of Bacillus strains in industrial bioreactors.
Exquisite Regioselectivity and Increased Transesterification Activity of an Immobilized Bacillus subtilis Protease
Authors: Ferreira Lino , Ramos M. A. , Gil Maria Helena , Dordick Jonathan S. 
Published: October 2002

AbstractCommercially available proteases and lipases were screened for their ability to acylate regioselectively sucrose with divinyladipate either in pyridine or dimethylformamide (DMF). The protease (EC 3.4.21.62) from Bacillus subtilis (Proleather FG‐F) exhibited the highest conversion (100% in 24 h of reaction in DMF) yielding sucrose 2‐ O‐vinyladipate as main product. The enzyme preference for a secondary hydroxyl group is a distinct feature of this biocatalyst compared to others described in the literature. Two sets of chemically distinct silica supports were used for Proleather immobilization presenting terminal amino (SAPTES) or hydroxyl groups (STESPM‐pHEMA). The percentage of immobilized enzyme was smaller in SAPTES (7–17%) than in STESPM‐pHEMA (52–56%), yet Proleather immobilized into SAPTES supports presented higher total and specific hydrolytic activity. The highest total and specific activities were obtained with STESPM‐pHEMA and SAPTES, respectively. Silicas with large pore (bimodal distribution of pores, 130/1200 Å, denoted as S1000) presented higher specific activities relative to those with smaller pore sizes. Furthermore, the synthetic specific activity of S1000SAPTES immobilized protease was ca. 10‐fold higher than that of the free enzyme. In addition to sucrose, the immobilized protease was used to acylate methyl α‐d‐glucopyranoside, trehalose, and maltose in nearly anhydrous DMF. Finally, immobilized Proleather was reasonably stable, retaining ca. 55% activity after six reaction cycles.
Continuous Selective Extraction of Secondary Metabolites from Catharanthus roseus Hairy Roots with Silicon Oil in a Two‐Liquid‐Phase Bioreactor
Authors: Tikhomiroff Cyril , Allais Ségolène , Klvana Maya , Hisiger Steve , Jolicoeur Mario 
Published: October 2002

AbstractA two‐liquid‐phase bioreactor was designed to extract indole alkaloids from Catharanthus roseus hairy roots with silicon oil. Partition studies between silicon oil and culture medium showed that the silicon oil did not alter the availability of nutrients. The affinity of tabersonine and löchnericine for silicon oil is nine times higher than for the aqueous phase. Cultures were elicited with 25 mg/L of jasmonic acid. The growth of the hairy roots was not significantly modified by the presence of silicon oil. The overall specific yields of tabersonine and löchnericine were increased by 100–400 % and 14–200 %, respectively, with the use of silicon oil in nonelicited control cultures. In elicited cultures, these values were 10–55 % for tabersonine and 20–65 % for löchnericine. Serpentine was never found in the silicon oil. All measured alkaloids' specific yields were higher using silicon oil and elicitation, suggesting that the silicon oil, while acting as a metabolic sink for tabersonine and löchnericine, was efficient in increasing metabolic fluxes of the secondary metabolism pathways.
Dual Excitation Ratiometric Fluorescent pH Sensor for Noninvasive Bioprocess Monitoring: Development and Application
Authors: Kermis Haley R. , Kostov Yordan , Harms Peter , Rao Govind 
Published: October 2002

AbstractThe development and application of a fluorescent excitation‐ratiometric, noninvasive pH sensor for continuous on‐line fermentation monitoring is presented. The ratiometric approach is robust and insensitive to factors such as source intensity, photobleaching, or orientation of the patch, and since measurements can be made with external instrumentation and without direct contact with the patch, detection is completely noninvasive. The fluorescent dye 8‐hydroxy‐1,3,6‐pyrene trisulfonic acid was immobilized onto Dowex strongly basic anion‐exchange resin, which was subsequently entrapped into a proton‐permeable hydrogel layer. The sensor layer was polymerized directly onto a white microfiltration membrane backing that provided an optical barrier to the fluorescence and scatter of the fermentation medium. The ratio of emission intensity at 515 nm excited at 468 nm to that excited at 408 nm correlated well with the pH of clear buffers, over the pH range of 6–9. The sensor responded rapidly (<9 min) and reversibly to changes in the solution pH with high precision. The sterilizable HPTS sensor was used for on‐line pH monitoring of an E. coli fermentation. The output from the indwelling sensor patch was always in good agreement with the pH recorded off‐line with an ISFET probe, with a maximum discrepancy of 0.05 pH units. The sensor is easily adaptable to closed‐loop feedback control systems.
Model‐Independent Relationships between Hematocrit, Blood Viscosity, and Yield Stress Derived from Couette Viscometry Data
Authors: Yeow Y. Leong , Wickramasinghe S. Ranil , Leong Yee‐Kwong , Han Binbing 
Published: October 2002

AbstractThis paper describes a procedure, based on Tikhonov regularization, for obtaining the shear rate function or equivalently the viscosity function of blood from Couette viscometry data. For data sets that include points where the sample in the annulus is partially sheared the yield stress of blood will also be obtained. For data sets that do not contain partially sheared points, provided the shear stress is sufficiently low, a different method of estimating the yield stress is proposed. Both the shear rate function and yield stress obtained in this investigation are independent of any rheological model of blood. This procedure is applied to a large set of Couette viscometer data taken from the literature. Results in the form of shear rate and viscosity functions and yield stress are presented for a wide range of hematocrits and are compared against those reported by the originators of the data and against independently measured shear properties of blood.
The Application of a Generic Feedstock from Wheat for Microbial Fermentations
Authors: Wang Ruohang , Domínguez‐Espinosa Rosa M. , Leonard Karen , Koutinas Apostolis , Webb Colin 
Published: October 2002

AbstractThe feasibility of a generic fermentation feedstock produced from wheat flour has been confirmed in several fermentations of yeasts, bacterium, and filamentous fungus for the production of commodity chemicals. Saccharomyces cerevisiae was incubated for the observation of yeast growth and ethanol production, Pichia farinosa for glycerol production, Monascus purpureus for fungal growth and pigment production, and Lactobacillus bulgaricus for bacteria growth and lactic acid production. The results confirmed that the feedstock contained no inhibitory components to the strains tested. Similar or higher metabolite yields were obtained in comparison with other studies carried in commonly used media.
Verbs of placement in Swedish children with SLI
Authors: Hansson Kristina , Bruce Barbro 
Published: October 2002

AbstractThis study investigated lexical aspects of verb knowledge in Swedish children with specific language impairment (SLI). The type of verbs studied were those of placement, a semantic area with a language‐specific differentiation in Swedish. It was hypothesized that owing to their cross‐linguistic uncommonness, these verbs cause difficulties for children with SLI, that these problems are more marked in production than in comprehension, and that children with SLI show less change over time than controls. Ten children with SLI and 10 language‐matched controls participated in an acting‐out task testing the comprehension and production of four different verbs of placement. The children were tested on two occasions, with a 6‐month interval. The groups did not differ in their performance on the comprehension task at either testing. In production, the children with SLI had significantly more difficulties in choosing the correct verb than the controls at the second testing. The controls performed significantly better on production at the second testing than at the first, whereas the children with SLI did not enhance their performance between testings. This indicates that children with SLI have lexical problems and that these problems do not diminish with time. The results are related to the results from studies of second‐language learners of Swedish who often have difficulties in the usage of these verbs. The results are also related to the results from studies of the corresponding semantic field in English‐ and Dutch‐speaking children with SLI. The comparisons show some interesting differences, depending on the type of learner and the language learned. The approach adopted seems promising. Studies of lexical specificity in children with SLI would give interesting information, useful both for the development of clinical assessment instruments and for our understanding of lexical acquisition in these children.
Cultural variations in attention regulation: A comparative analysis of British and Chinese‐immigrant populations
Authors: Vigil Debra C. 
Published: October 2002

AbstractRecent intervention for young children has included parent training. The premise for this training is based in research on parent‐child interaction. However, these studies have focused on white, middle‐class families. More recent research suggests that not all cultures follow the same pattern. This study explored the relationship between attention regulation, pragmatic input, object manipulation and later language competence in Chinese‐immigrant and British caregiver‐infant dyads. Microanalytical techniques were used to explore observational data derived from a longitudinal study of 10 caregivers and their infants at 9 and 12 months of age. Relations among selected mother and infant activities, notably caregiver control of and responsiveness to attentional focus in infants and object/infant manipulation, were evaluated. Two styles of attention regulation were found across groups. British caretakers responded to their infant's behaviours by following their lead, whereas Chinese caretakers directed their infant's attention. These styles were supported by non‐verbal behaviours such as object manipulation on the part of the caretaker. No differences between the two groups in expressive vocabulary size at age 18 months were found, suggesting that attention regulation styles of directing a child's attention or following a child's lead need further investigation to determine the relationship with vocabulary acquisition. Results show that parents begin to use a particular style of interaction with their children at a very young age. Further research is required to determine if this style continues through the language‐learning years. Furthermore, this study suggests that speech and language therapists must be aware that the communication style of ethnic minorities on their caseloads maybe different from mainstream culture; and with this knowledge service providers will be able to serve those communities better and avoid potential cultural bias.
Mass Spectrometric Identification of Isoforms of PR Proteins in Xylem Sap of Fungus-Infected Tomato
Authors: Rep Martijn , Dekker Henk L. , Vossen Jack H. , de Boer Albert D. , Houterman Petra M. , Speijer Dave , Back Jaap W. , de Koster Chris G. , Cornelissen Ben J.C. 
Published: October 2002

Abstract
The protein content of tomato (Lycopersicon esculentum) xylem sap was found to change dramatically upon infection with the vascular wilt fungus Fusarium oxysporum. Peptide mass fingerprinting and mass spectrometric sequencing were used to identify the most abundant proteins appearing during compatible or incompatible interactions. A new member of the PR-5 family was identified that accumulated early in both types of interaction. Other pathogenesis-related proteins appeared in compatible interactions only, concomitantly with disease development. This study demonstrates the feasibility of using proteomics for the identification of known and novel proteins in xylem sap, and provides insights into plant-pathogen interactions in vascular wilt diseases.
Transient Induction of Cyclin T1 during Human Macrophage Differentiation Regulates Human Immunodeficiency Virus Type 1 Tat Transactivation Function
Authors: Liou Li-Ying , Herrmann Christine H. , Rice Andrew P. 
Published: October 2002

ABSTRACTThe human immunodeficiency virus type 1 (HIV-1) Tat protein is essential for viral replication and stimulates transcription of the integrated provirus by recruiting the kinase complex TAK/P-TEFb, composed of cyclin T1 (CycT1) and Cdk9, to the viral TAR RNA element. TAK/P-TEFb phosphorylates the RNA polymerase II complex and stimulates transcriptional elongation. In this report, we investigated the regulation of TAK/P-TEFb in primary human macrophages, a major target cell of HIV infection. While Cdk9 levels remained constant, CycT1 protein expression in freshly isolated monocytes was very low, increased early during macrophage differentiation, and, unexpectedly, decreased to very low levels after about 1 week in culture. The kinase activity of TAK/P-TEFb paralleled the changes in CycT1 protein expression. RNA analysis indicated that the transient induction of CycT1 protein expression involves a posttranscriptional mechanism. In transient transfection assays, the ability of Tat to transactivate the HIV long terminal repeat (LTR) in the late differentiated macrophages was greatly diminished relative to its ability to transactivate the HIV LTR in early differentiated cells, strongly suggesting that CycT1 is limiting for Tat function in late differentiated macrophages. Interestingly, lipopolysaccharide, a component of the cell wall of gram-negative bacteria, reinduced CycT1 expression late in macrophage differentiation. These results raise the possibility that regulation of CycT1 expression may be involved in establishing latent infection in macrophages and that opportunistic infection may reactivate the virus by inducing CycT1 expression.