摘要:

AbstractA structured metabolic model is developed that describes the stoichiometry and kinetics of the biological P removal process. In this approach all relevant metabolic reactions underlying the metabolism, considering also components like adenosine triphosphate (ATP) and nic‐otinamide‐adenine dinucleotide (NADH2) are describedbased on biochemical pathways. As a consequence of the relations between the stoichiometry of the metabolic reactions and the reaction rates of components, the required number of kinetic relations to describe the process is reduced. The model describes the dynamics of the storage compounds which are considered separately from the active biomass. The model was validated in experiments at a constant sludge retention time of 8 days, over the anaerobic and aerobic phases in which the external oncentrations as well as the internal fractions of the relevant components involved in the P‐removal process were monitored. These measurements include dissolved acetate, phosphate, and ammonium; oxygen consumption; poly‐β‐hydroxybutyrate (PHB); glycogen; and active biomass. The model satisfactorily describes the dynamic behavior of all components during the anaerobicand aerobic phases.© 1995 John Wil

ISSN:0006-3592    

DOI:10.1002/bit.260470302

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractPeptide libraries can be used to identify ligands that bind specifically to a desired protein. These peptides may have significant advantages as specific ligands for affinity chromatography separations. This article describes the use of one of such peptide, Try‐Asn‐Phe‐Glu‐Val‐Leu, as a ligand for the purification of S‐protein using affinity chromatography. General strategies for peptide immobilization are discussed and the conditions for peptide immobilization to Emphaze™ gel are optimized. The effects of peptide orientation and peptide densities on protein binding are studied. Results indicate that the peptide affinity is not affected by the orientation of the peptide during immobilization, but association constants can be reduced by one order of magnitude when compared with the values in solution.With increased peptide density, the protein binding capacity of the gel increases, but both the percentage of peptide utilization and apparent binding constant between immobilized peptide and S‐protein decrease. S‐protein is separated from a mixture with BSA via affinity chromatography using specific elution with the peptide in solution.Finally, direct purification of S‐protein from an enzymatic digestion mixture of ribonuclease A is demonstrated.© 1995

ISSN:0006-3592    

DOI:10.1002/bit.260470303

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractThe simultaneous biodegradation of toxic compounds in mixtures is a major current concern. To bioremediate a toxic mixture, we designed a strategy combining an ad‐sorbent carrier with an ecological and nutritional system which allowed work close to heavily polluted conditions in nature. Starting from a methanogenic community, we developed a microbial consortium acclimated to a mixture of about 30 chlorinated aliphatics in a fixed‐film stationary‐bed bioreactor. Prior to the establishment of a durable period of dechlorination, an interval of progressive dechlorination of the toxic mixture was observed during which the excess of the toxic compounds was stored on the carrier. The latter, consisting of activated carbon in a polyurethane foam, allowed us to work at concentrations far above the solubility of the toxic compounds (apparent concentrations of about 10 g/L). The complete disappearance of hexachloroethane as well as its lower homologues, penta‐, tetra‐, and trichloroethane, present in the toxic mixture, was observed. Additionally, octachlorocyclopentene, carbon tetrachloride, trichloro‐ethylene, tetrachloroethylene, and hexachloro‐1,3‐butadiene also completely disappeared. For the four latter compounds, from mass balances in the bioreactor, degradation rates around 10 μmol/L per day were determined with total dechlorination. The enrichment culture thus developed exhibited high degradation performances similar to those reported in the literature for pure or enriched anaerobic microbial cultures in contact with a single toxic compound. The results demonstrate the possibility of concurrent high‐rate degradation of several highly chlorinated toxic compounds, under conditions approximating field situations.© 1995

ISSN:0006-3592    

DOI:10.1002/bit.260470304

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractAmmonium and lactate are two known toxic products detrimental to mammalian cell growth and productivity. An electrokinetic technique, utilizing an electrophoretic mechanism, was developed to remove these cellular wastes in‐situ from suspension hybridoma (ATCC CRL‐1606) cultures to enhance cell growth and productivity. This technique applies continuously a dc electric field to selectively remove the electrically charged wastes. The experiments were shown to be successful in the removal of externally added 10 rnMammonium and 45 mMlactate while maintaining the chemostatic condition of culture medium in a cell‐free condition under an electric current density of 50 A/m2. Toxic levels of ammonium were added, ranging from 7.5 to 12.5 mM, at the start of the hybridoma culture, and the applied dc electric fields were able to completely remove these added materials. This in turn released the inhibition and restored the cell growth. Finally, this electrokinetic technique was applied to the batch and glutamine fed‐batch hybridoma cultures. At an applied electric current density of 50 A/m2, this was able to completely remove cell‐produced ammonium and increased the cell growth and antibody titer by 30% to 50%, respectively, compared to the control experiment in the absence of the electric field. Lastly, the applied electric current density of 50 A/m2did not affect cellular functionalities such as glucose and glutamine consumption and antibody productivity.© 1995 John Wile

ISSN:0006-3592    

DOI:10.1002/bit.260470305

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractIn‐situ dc electric fields were applied to remove ammonium and lactate from suspension hybridoma cultures (ATCC‐CRL‐1606) which used enriched media. Nutrient concentration was increased fourfold above the normal concentration of DMEM to study enhanced protein product formation in a dc electric field. In the presence of the electric field, hybridoma growth and antibody production were increased 1.5‐fold (from 3.7 × 106to 9.1 × 106viable cells/mL) and twofold (from 170 to 505 mg IgG/L), respectively, compared with the control. The effective removal of ammonium and lactate and increased concentrations of the various nutrients accounted for this enhancement. The enriched media caused the overflow metabolism of glucose, glutamine, and various essential amino acids. The overconsumption of glucose also produced substantial amounts of lactate, which in turn greatly increased the medium osmolarity. The increase in medium osmolarity is believed to be one of the causes of cell death in these culture systems.© 1995 John Wile

ISSN:0006-3592    

DOI:10.1002/bit.260470306

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractThis study shows that, in a sulphide‐oxidizing bioreactor with a mixed culture ofThiobacilli, the formation of sulphur and sulphate as end‐products from the oxidation of sulphide can be controiledinstantaneously and reversibiy by the amount of oxygen supplied. It was found that at sulphide loading rates of up to 2.33 mmol7/L · h, both products can be formed already at oxygen concentrations below 0.1 mg/L. Because the microorganisms tend to form sulphate rather than forming sulphur, the oxygen concentration is not appropriate to optimize the sulphur production. Within less than 2 h, the system can be switched reversibly from sulphur to sulphate formation by adjusting the oxygen flow. This is below the minimum doubling time (2.85 h) of, e.g.,Thiobacillus neapolitanusandThiobacillus 0,18which indicates that one metabolic type of organism can probably perform both reactions. Under highly oxygen‐limited circumstances, that is, at an oxygen/sulphide consumption ratio below 0.7 mol · h−1mol · h−1thiosulphate is abundantly formed. Because the chemical sulphide oxidation results mainly in the formation of thiosulphate, it is concluded that, under these circumstances, the biological oxidation capacity of the system is lower than the chemical oxidation capacity. The oxidation rate of the chemical sulphide oxidation can be described by a first‐order process (k=−0.87 h−1).© 1995

ISSN:0006-3592    

DOI:10.1002/bit.260470307

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractA number of factors have been shown to affect the metabolism of glucose and glutamine in mammalian cells and their mechanisms have been partially elucidated. Despite these efforts, a quantitative knowledge of the significance of these factors, the regulation of glucose and glutamine utilization, and particularly the interactions of these two nutrients is still lacking. Controversies exist in the literature. To clarify some of these controversies, mathematical models are proposed in this work which enable to separate and identify the effects of individual factors. Experimental data from five cell lines obtained in batch, fed‐batch, and continuous cultures, both under steady‐state and transient conditions, were used to verify the model formulations. The resulting kinetic models successfully describe all these cultures. According to the models, the specific consumption rate of glucose (QGlc) of continuous animal cells under normal culture conditions can be expressed as a sum of three parts: a part owing to cell growth; a part owing to glucose excess; and a part owing to glutamine regulation. The specific consumption rate of glutamine (qGlc7) can be expressed as a sum of only two parts: a part owing to cell growth; and a part owing to glutamine excess. Using the kinetic models the interaction and regulation of glucose and glutamine utilizations are quantitatively analyzed. The results indicate that, whereasqGlcis affected by glutamine,qGlnappears to be not or less significantly affected by glucose. It is also shown that the relative utilizations of glucose and glutamine by anabolism and catabolism are mainly affected by the residual concentrations of the respective compounds and are less sensitive to growth rate and the nature of growth limitation.© 1995 John Wiley&Sons

ISSN:0006-3592    

DOI:10.1002/bit.260470308

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractOn‐line NAD(P)H fluorescence and culture redox potential (CRP) measurements were utilized to investigate the role ofVitreoscillahemoglobin (VHb) in perturbing oxygen metabolism of microaerobicEscherichia coliBatch cultures of a VHb‐synthesizingE. colistrain 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 keepsE. coliin 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.© Jo

ISSN:0006-3592    

DOI:10.1002/bit.260470309

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractNonlinear adsorption plays an important role in determining the chromatographic behavior of proteins in preparative ion‐exchange chromatography. In this article, the steric mass action (SMA) isotherm is used in conjunction with a mass transport model to describe nonlinear cation‐exchange chromatography. Excellent agreement is observed between simulated and experimental step gradient separations of the proteins α‐chymotryp‐sinogen A, cytochromeC, and lysozyme. A systematic method of selecting the optimum step gradient program for a given separation problem is presented and employed to study optimization of step gradient chromatography under conditions of high mass loading. This article includes consideration of the effects of the adsorption properties of the feed stream, the feed stream concentration, protein solubility, and otherconstraints on the optimum separation conditions.© John Wiley&

ISSN:0006-3592    

DOI:10.1002/bit.260470310

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractThis study evaluated the feasibility of monitoring nitro‐genase activity in situ through measurement of N2uptake rate (NUR) using off‐gas mass spectrometry. Four 50‐L cultures ofAzotobacter vinelandiiwere grown aer‐obically in nitrogen‐free medium to cell densities of 1.0‐1.3gL−1magnetic‐sector mass spectrometer was used to monitor NUR along with other gas exchange rates. The small specific uptake rate (1.2 mmol g−1h−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% N215% Ar) were designed to improve the signal‐to‐noise ratio while maintaining dissolved O2and N2levels in desired ranges. The fraction of N2removed 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−1h−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

ISSN:0006-3592    

DOI:10.1002/bit.260470311

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY