bluuebunny/crossref_metadata_2025_split huggingface dataset:
The influence of bond chemistry on immobilized enzyme systems for ex vivo use
Authors: Comfort Ann R. , Mullon Claudy J.‐P. , Langer Robert 
Published: December 2004

AbstractThe ideal derivatized support for the clinical use of an immobilized enzyme system should irreversibly bind active enzyme. We have investigated the behavior of heparinase and bilirubin oxidase immobilized via cyanogen bromide, tresyl chloride, epoxide, or carbodiimidazole activated natural and synthetic matrices. The protein bound to each activated support was 90% for cyanogen bromide (CNBr) activated agarose, 50–80% for tresyl chloride activated agarose, and 50% for oxirane activated acrylic (Eupergit C). The activity retention of immobilized heparinase was greatest (50%) with CNBr activated agarose while for the immobilization of bilirubin oxidase, the activity retention was greatest (25–30%) with tresyl chloride activated agarose and oxirane activated acrylic.The stability of the different covalent bonds was studied in vitro with radioiodinated enzymes. The leaching profiles showed the same trends for each support and chemistry. A plateau in portein leaching was reached after a few hours of incubatttion and the transient leaching period was well represented byu a logarithimic function of time. The amount of enzyme released from the least stable support (CNBr activated agarose) in 24 h was injected intravenously in New Zealand white rabbits. Using an indirect enzyme‐linked immunnosorbant assay (ELISA), no immune responce was detected. The transient leaching profile was shortenend by washingthe enzyme‐support conjugate with 1M hydroxylamine, pH8.5 intermolecular cross‐linking with glutaraldehyde also improves the enzyme‐support stability. Tresyl chloride and oxirane activated supports produce bonds with improved stability without adversely affecting enzymatic activity.
Use of polyelectrolyte complex‐stabilized calcium alginate gel for entrapment of β‐amylase
Authors: Kokufuta Etsuo , Shimizu Noboru , Tanaka Hideo , Nakamura Isei 
Published: December 2004

AbstractCalcium alginate gel stabilized with a polyelectrolyte complex (PEC) consisting of potassium poly(vinyl alcohol) sulfate (KPVS) and trimethylammonium glycol chitosan iodide (TGCI) was used for the immobilization of β‐amylase. The immobilization was made by gelling aqueous droplets of enzyme solution including both sodium alginate and KPVS in a CaCl2 solution containing TGCI. The activity of the enzyme entrapped into the stabilized gel beads was evaluated by studying the batch reaction kinetics of enzyme‐catalyzed hydrolysis of maltotetraose. Repeated kinetic measurements, totaling 18, were carried out at fixed time intervals. After each measurement the beads were stirred for 1 day in a freshly prepared 10 mM NaCl solution at 3°C. It was found that the immobilized system remained stable without leading to a serious loss of the activity or to a large leakage of the enzyme from the support. This was explained as being due to a PEC‐crosslinked contracted network structure of the stabilized gel matrix.
Solventogenesis in Clostridium acetobutylicum fermentations related to carboxylic acid and proton concentrations
Authors: Hüsemann Michael H. W. , Papoutsakis Eleftherios T. 
Published: December 2004

AbstractThe mechanism primarily implicated in the solventogenesis process in batch fermentations of Clostridium acetobutylicum is examined in considerable detail. A variety of fermentations with or without pH control in the pH range of 3.7–6 have been carried out in order to examine which of a host of suspect parameters correlate with the initiation of solventogenesis. The parameters that did not correlate are the external (pH0) and intracellular (pHi) pH, and ΔpH, and the external or intracellular butyrate and acetate concentrations. Undissociated butyric acid (UBA) correlated well with the initiation of solventogenesis. A linear relationship between UBA and butanol concentrations was found at the onset of solventogenesis in all fermentations examined. The intercept of this linear relationship was 6–13mM UBA for the pH0 range of 3.7–5 and approximately zero for pH0 at or above 6. The required minimal UBA was interpreted as a dependency of the solventogenesis process on both H+ and butyrate concentrations. Undissociated acetic acid was found not to correlate with the initiation of solventogenesis. Addition of acetoacetate (AA) and propionate enhanced the effect of UBA on the solventogenesis process. The action of a nonmetabolizable (FCCP) and a metabolizable (AA) uncoupler on the ΔpH, pH0, pHi, and solventogenesis were also studied in order to gain further understanding of the solventogenesis mechanism.
Removal of lignin and reuse of cellulases for continuous saccharification of lignocelluloses
Authors: Tanaka Mitsuo , Fukui Masahide , Matsuno Ryuichi 
Published: December 2004

AbstractMethod of the removal of lignin and reuse of cellulases for a continuous saccharification of lignocelluloses were investigated. Only lignin could be separated from hydrolysates by differences in the settling velocity; it was removed from the saccharification process by flocculation with chitosan without loss of cellulases. The ultra‐filtration membrane PM10 (Amicon) could be used for recovery of cellulases, but the membrane UH‐1 (Toyo Roshi) was better for this purpose, because no cellulases leaked from the membrane, and the amount of cellulase adsorbed to the membrane was less. The cellulases were inactivated by vigorous agitation of the solution in an ultra‐filtration device. The loss of cellulase activity by such agitation increased with agitation time, but could be controlled by recovery at a low speed of agitation, so the cellulases could be reused.
Continuous processing of toxic organics in a fluidized‐bed GAC reactor employing carbon replacement
Authors: Pfeffer John T. , Suidan Makram T. 
Published: December 2004

AbstractThe fluidized‐bed, granular activated carbon (GAC) anaerobic reactor has been shown to be an effective process for the continuous long‐term treatment of wastewaters that contain biodegradable or nonbiodegradable toxic organic compounds. With loadings of 10 g COD/kg GAC day, COD removal of 94% was achieved. The anaerobic biofilm that develops on the GAC reduces the load on the carbon by converting the biodegradable organics to methane and carbon dioxide. Approximately 50% of the COD applied to the reactor was converted to methane, thereby reducing carbon requirements. Successful operation of the system requires that a carbon replacement schedule be maintained that will keep the bulk concentrations of toxic adsorbable compounds below their toxic threshold. As long as toxic substances can be adsorbed by the carbon, they will not inhibit the anaerobic biofilm. If nonadsorbable toxic compounds are present, processing must be included to reduce these materials to concentrations below their threshold level.
Mixed substrate‐metabolism in an obligate methanol utilizer: Chemostat studies
Authors: Samuelov Nissim S. 
Published: December 2004

AbstractIn Pseudomonas C, an obligate RMP‐methylotroph, externally added formic acid and multi‐carbon intermediates are co‐metabolized in the obligate presence of methanol. Cell yield and methanol‐carbon conversion to cell mass are optimal in a chemostat culture growing on a mixture of energy sources and methanol as the sole limiting carbon source. Growth on methanol as the sole, obligate energy source and an additional carbon source (e.g. pyruvate) is less efficient. Under these energy limited conditions, the cyclic oxidation pathway is predominantly operated.
A model for continuous fermentations with amylolytic yeasts
Authors: Pasari Ajay B. , Korus Roger A. , Heimsch Richard C. 
Published: December 2004

AbstractA two‐stage, associative fermentation process is more effective for continuous yeast biomass production from starch than a single‐stage mixed culture fermentation process. By operating two stages, competition for the same growth limiting substrate is reduced leading to efficient starch utilization. In this article, a mathematical model has been proposed for continuous, two‐stage fermentation with a pure culture, amylolytic yeast in the first stage and a mixed culture second stage with a faster growing, nonamylolytic yeast. The model parameters were determined for Saccharomycopsis fibuligera and Candida utilis in continuous, single‐stage, pure cultures. In the two‐stage model, the effects of changes in dilution rate on biomass, amylase, reducing sugar, and starch concentration, and ratio of stage volumes on microbial composition are discussed and compared with experimental data.
An algorithmic approach to constructing the on‐line estimation system for the specific growth rate
Authors: Shimizu Hiroshi , Takamatsu Takeichiro , Shioya Suteaki , Suga Ken‐Ichi 
Published: December 2004

AbstractThe objective of this article is to propose an algorithm for the on‐line estimation of the specific growth rate in a batch or a fed‐batch fermentation process. The algorithm shows the practical procedure for the estimation method utilizing the macroscopic balance and the extended Kalman filter. A number of studies of the on line estimation have been presented. However, there are few studies discussing about the selection of the observed variables and for the tuning of some parameters of the extended Kalman filter, such as covariance matrix and initial values of the state.The beginning of this article is devoted to explain the selection of the observed variable. This information is very important in terms of the practical know‐how for using technique. It is discovered that the condition number is a practically useful and valid criterion for number is a practically useful and valid criterion for choosing the variable to be observed.Next, when the extended Kalman filter in applied to the online estimation of the specific growth rate, which is directly unmeasurable, criteria for judging the validity of the estimated value from the observed data are proposed. Based on the proposed criterial, the system equation of the specific growth rate is selected and initial value of the state variable and covariance matrix of the system noises are adjusted. From many experiments, it is certified that the specific growth rate in the batch or fed ‐batch fermentation can be estimated accurately by means of the algorithm proposed here. In these experiments, that is, when the cell concentration is measured directly, the extended Kalman filter using the convariance matrix with a constant element can estimate more accurately values of the specific growth rate than the adaptive extended Kalman filter does.
Multiple forms of endo‐1,4‐β‐D‐glucanase in the extracellular cellulase of Penicillium pinophilum
Authors: Bhat K. Mahalingeshwara , Wood Thomas M. 
Published: December 2004

AbstractThe influence of the composition of the growth medium on the production of endo‐1,4‐β‐D‐glucanase (CM‐cellulase) activity by P. pinophilum was studied in shake flask cultures using Avicel PH101 as the carbon source. It was observed that the culture conditions had a profound effect on the level of endoglucanase (CM‐cellulase) produced by P. pinophilum. However, isoelectric focusing of the endoglucanase activity obtained from shake flask and fermenter cultures using the same growth medium revealed that the enzyme system found in both cultures was identical qualitatively, and contained seven or eight different endoglucanase components. All the endoglucanase components appeared simultaneously in the early stages of culture and prolonged incubation resulted only in an increase in the concentration of these enzymes. Protease levels were found to be low in both types of culture but were particularly so in the growth medium which contained corn steep liquor. The proteases were unable to release low molecular weight peptides when P. pinophilum cellulase protein was used as a substrate. The results were interpreted to indicate that the multiplicity of endoglucanase components found in cultures of P. pinophilum is most likely the result of expression of a number of specific genes rather than by post‐secretional modification of one or more endoglucanase(s) synthesized by the fungus.
Water‐soluble nonionic surfactants for affinity bioseparations
Authors: Guzman R. , Torres J. L. , Carbonell R. G. , Kilpatrick P. K. 
Published: December 2004

AbstractReversible competitive inhibitors of the three enzymes β‐galactosidase, trypsin, and serum cholinesterase have been covalently attached to nonionic ethoxylated surfactants. The binding of the resulting affinity‐derivatized surfactants to the respective enzymes has been quantified by measuring Michaelis‐Menten inhibition constants with kinetic assays. The surfactant‐inhibitor of serum cholinesterase, octaethylene glycol monohexadecy ether pyridinium (C16E8‐PYR), was adsorbed in aqueous solution to an octadecyl‐bonded reverse‐phase silica packing in a 2 × 0.2 cm stainless steel test column. The ability of the test column to function as a high‐performance affinity chromatography (HPAC) column was determined by applying a mixture of bovine serum albumin and cholinesterase (4:1 w/w). Virtually all of the cholinesterase bound and was eluted by applying a gradient in ionic strength. The applied cholinfesterase was recovered with a yield of over 90% and an 11‐fold purification. An aliquot of raw horse serum was then purified in the same fashion with a yield of 84% and a 280‐fold purification. The surfactant‐inhibitor was easily removed from the column with an alcohol wash for sterilization, cleaning, or application of a different affinity ligand. Moreover, the ligand density on the column can be easily manipulated by adsorbing mixtures of derivatized and underivatized surfactants. Leakage of ligands from the support seems to be minimal since the cholinesterase affinity column was operated efficiently after being exposed to 24,000 column volumes of buffer. The application of this technique to high‐capacity, high‐throughput reversible affinity purifications is limited only by the ability to identify suitable ligands.
Degradation of fluorene in soil by fungus Phanerochaete chrysosporium
Authors: George Elias J. , Neufeld Ronald D. 
Published: December 2004

AbstractDuring investigation of biodegradation in soil, we have found that classical or standard techniques for introduction of compounds and the growth of fungus into soil are ill‐defined and inadequate. In response to this deficiency, a method for controlled introduction of extractable compounds and for the growth of fungus in soils has been developed. This method was successfully used to study the degradation of fluorene in soil by the fungus Phanerochaete chrysosporium.
Stabilization of restriction endonuclease Bam HI by cross‐linking reagents
Authors: Dubey A. K. , Bisaria V. S. , Mukhopadhyay S. N. , Ghose T. K. 
Published: December 2004

AbstractBacillus amyloliquefaciens H produces a restriction endonuclease enzyme BamHl which is heat labile even at low temperatures. Studies were conducted to enhance thermal stability of BamHl using cross‐linking reagents, namely, glutaraldehyde, dimethyl adipimidate (DMA), dimethyl suberimidate (DMS), and dimethyl 3,3′‐dithiobispropionimidate (DTBP). Reaction with glutaraldehyde did not result in a preparation with enhanced thermal stability. However, the DMA‐, DMS‐, and DTBP‐cross‐linked preparations of BamHI exhibited significant improvement in thermal stability. Studies on thermal denaturation of the cross‐linked enzyme preparations revealed that these do not follow a true first‐order kinetics A possible deactivation scheme has been proposed in which the enzyme has been envisaged to go through a fully active but more susceptible transient state which, on prolonged heat exposure, exhibits a first‐order decay kinetics. At 35°C, which is close to the optimum reaction temperature of 37°C for BamHl activity, the half‐line of DMA‐, DMS‐, and DTBP‐cross‐linked preparations were 4.0, 5.25, and 5.5 h, respectively, whereas the native enzyme exhibited a half‐line of 1.2 h only. The apparent values of deactivation rate constants for native, DMA‐, DMS‐, and DTBP‐cross‐linked BamHl were 1.13, 0.39, 0.29, and 0.26 h−1, respectively, at the same temperature, and the apparent values of activation energies for denaturation of native, DMA‐, DMS‐, and DTBP‐cross‐linked BamHl were 2.63, 5.24, 6.55, and 9.2 kcal/mol, respectively. The DTBP‐cross‐linked Bam HI was, therefore, the best heat‐stable preparation among those tested. The unusually low values of activation energies for denaturation of Bam Hl represent their highly thermolabile nature compared to other commonly encountered enzymes such as trypsin, having activation energies of more than 40 kcal/mol for their denaturation.
The effect of concentration of phenols on their batch methanogenesis
Authors: Wang Yi‐Tin , Suidan Makram T. , Pfeffer John T. , Najam Issam 
Published: December 2004

AbstractThe biodegradability of phenol and six other phenolic compounds (o‐, m‐, and p‐cresol, 2‐, 3‐, and 4‐ethylphenol) was examined in batch methanogenic cultures. The effect of concentration of these alkyl phenols on the anaerobic biodegradation of phenol was also evaluated. The inoculum used in this study was cultivated in a continuous flow laboratory fermenter with phenol as the primary substrate. Phenol, at initial concentrations as high to 1400 mg/L was completely degraded to methane and carbondioxide after 350 hours incubation. Complete degradation of m‐ and p‐cresol was also observed while the ethylphenols and o‐cresol were not significantly degraded.At initial concentrations exceeding 600 mg/L, phenol inhibited the phenol‐degrading microorganisms but not the methanogens. At about 600 mg/L, cresols reduced the rate of phenol degradation to 50% of that observed in a control culture containing only 200 mg/L phenol. Ethylphenols were more inhibitory than cresols. Phenol degrading microorganisms were more susceptible to inhibition by cresols and ethylphenols than were the methanogens. The inhibitory effects of the three isomers of cresol and ethylphenol did not vary with the isomer but rather with the substituted functional group.
On‐line monitoring of cell mass in mammalian cell cultures by acoustic densitometry
Authors: Kilburn D. G. , Fitzpatrick P. , Blake‐Coleman B. C. , Clarke D. J. , Griffiths J. B. 
Published: December 2004

AbstractAcoustic resonance densitometry (ARD) provides a highly reproducible and stable method for on‐line measurement of culture biomass density. The technique provides a direct determination of changes in relative density of culture medium and cell mass. At cell concentrations higher than 106 cells mL−1this method can replace cell counts and provide a continuous measure of total cell mass. In cultures of hybridomas or U937 human lymphoma cells, the ARD value correlates well with cell number except when the average cell size changes during culture. It is argued that cell mass determined by ARD rather than cell number should be used as the basis for measurements of specific biological activity.
Two‐Level factorial screening of new plasmid/strain combinations for prodution of recombinant‐DNA products
Authors: Emborg Claus , Jepsen Pia Knak , Biedermann Kirsten 
Published: December 2004

AbstractThis article treats the basic problem of selection of experimental conditions for microbiological experiments for evaluation of newly isolated bacterial strains, mutants, or plasmid/strain combinations. For this purpose shake flask experiments in a 210‐4confounded factorial design at resolution IV with four blocks of 16 flasks were used. The design was used for testing of two new strain/plasmid combinations (E. coli MT 102/403‐SD2 and W 3110/403‐SD2) i.e., both strains with the same plasmid 403‐SD2. Both strains were integrated in the design, so both strains were tested with nine factors (temperature, aeration, glucose, initial pH, pH regulation, reduced aeration, chloramphenicol, acetate, and glycerol). With both strains the interaction between initial pH and reduced aeration had a significant influence on the yield of the recombinant‐DNA product nuclease. There was more than a factor of 10 between lowest and highest yield of product. In this interactive system the strains reacted differently. MT 102/403‐SD2 had highest yields at high initial pH (8.4) and no reduction in aeration, whereas W 3110/403‐SD2 had highest yields of nuclease at low initial pH (7.4) and reduced aeration (rubber stopper inserted after cultivation for 12 h). These data (and previous work) clearly demonstrate that it is impossible to suggest a simple set of experimental conditions for testing of new plasmid/strain combinations. It is clear that the exclusive application of a standardized growth technique e.g., LB‐medium at 37°C at an unspecified and uncontrolled aeration level, may lead to wrong conclusions on properties and potentials of now plasmid/strain combinations and may lead to rejection of useful strains or plasmids.
The degradation of lignocellulosics by extremely thermophilic microorganisms
Authors: Donnison A. M. , Brockelsby C. M. , Morgan H. W. , Daniel R. M. 
Published: December 2004

AbstractFive extremely thermophilic cellulose‐degrading isolates obtained from New Zealand thermal springs were tested for their ability to degrade a number of natural lignocellulosic substrates. Degradation by three of the isolates was generally similar to that by the moderate thermophile, Clostridium thermocellum but occurred at a higher temperature. The New Zealand isolates were also found to grow on xylan as sole carbohydrate source, which probably extends their attack to hemicellulose.
Role of some whey components on mass transfer in ultrafiltration
Authors: Taddei C. , Daufin G. , Aimar P. , Sanchez V. 
Published: December 2004

AbstractUltrafiltration through Carbosep M4 mineral membrane of protein solutions of decreasing complexity (whey before and after centrifugation or clarification, β‐lactoglobulin) was studied. Mathematical models were used to explain variations in flux with time. Taking into account variations in protein retention and hydraulic resistance of the membrane during ultrafiltration, proteins and lipoproteins were found to be involved not only in the polarization layer (reversible fouling leading to a difference in the osmotic pressure), but also in irreversible fouling by adsorption. Morever, the presence of particles (e.g., inorganic precipitates) in whey explains the build‐up of a deposit over and within the membrane which contributes to the decline in flux after 1 h ultrafiltration. The relative importance of these phenomena was quantified.
An approach in mathematical modeling of an upflow packed‐bed reactor for enzymatic hydrolysis of wheat straw
Authors: Gonzàlez G. , Caminal G. , de Mas C. , Lòpez‐Santín J. 
Published: December 2004

AbstractThe behavior of a packed‐bed reactor for enzymatic hydrolysis of pretreated wheat straw has been described by means of a mathematical model. The flow pattern has been evaluated by residence time distribution experiments. Small deviations from ideal plug flow behavior were found using the dispersion model. The kinetic model proposed for enzymatic hydrolysis of cellulosic fraction of pretreated wheat straw has been derived from batch experimental data. Variations of enzyme concentration throughout the straw bed have been approximately described using a ramp variation of adsorbed enzyme. The final explains qualitatively the experimental results.
Lactose hydrolysis by immobilized β‐galactosidase in capillary bed reactor
Authors: Peterson R. S. , Hill C. G. , Amundson C. H. 
Published: December 2004

AbstractThe hydrolysis of lactose by immobilized β‐galactosidase was studied in a continuous‐flow capillary bed reactor operating at 30°C. Solutions containing 50, 100, and 150 g lactose and 0.5 g sodium acetate/L were fed to the reactor. Lactose conversions ranging from 24% to greater than 99% were achieved at reactor space times ranging from 0.06 to 6.3 min. These conversion data were successfully modeled in terms of a plug flow reactor model and a form of Michaelis–Menten kinetics which included competitive inhibition by both the alpha and beta forms of galactose.