bluuebunny/crossref_metadata_2025 huggingface dataset:
Determination of cellular rate distributions in microbial cell populations: Feeding rates of ciliated protozoa
Authors: Hatzis Christos , Sweeney Pamela J. , Srienc Friedrich , Fredrickson A. G.
Published: December 2004

A novel procedure is proposed for determining distributions of rate properties and correlations of rate with state properties of microbial cell populations. The procedure is novel in that it uses transient data, and thus, it does not require that the population be in balanced growth, although it requires that the population structure does not change during the short transient experiment. The procedure is applied to populations of the ciliated protozoan Tetrahymena to determine ingestion rate variability. The number of ingested microspheres per cell and the single‐cell protein content—an indicator of cell size—were directly determined with dual‐color flow cytometry. The proposed technique revealed the correlation pattern of the particle ingestion rate with cell size. In particular, ingestion rate was found to be positively correlated with cell size for the smaller feeding cells and to be uncorrelated with size for the larger cells. Using the fact that particle uptake from dilute particle suspensions is a Poisson random process, we determined that the coefficient of variation of the distribution of ingestion rates within the feeding population is about 50%. It was concluded that the dynamics of particle ingestion can be accurately described only if it is realized that particle ingestion rates are distributed. © 1993 John Wiley & Sons, Inc.
Synthesis of fatty acid esters by a recombinant cutinase in reversed micelles
Authors: Sebastião M. J. , Cabral J. M. S. , Aires‐Barros M. R.
Published: December 2004

Fusarium solani pisi recombinant cutinase, solubilized in AOT/isooctane‐reversed micelles, was used to catalyze the esterification of fatty acids with aliphatic alcohols. Some relevant parameters for the enzyme activity such as pH, Wo (water/surfactant molar ratio), temperature, and substrate concentration were optimized. Maximal specific activity was obtained for hexanol. The cutinase showed selectivity for short‐chain fatty acids. The stability of the microencapsulated cutinase was investigated at various concentrations of water and different values of pH. Oleic acid had a negative effect on the cutinase stability, while hexanol proved to be a strong stabilizer increasing the half‐life of the enzyme about 45 times. © 1993 John Wiley & Sons, Inc.
Charged fusions for selective recovery of β‐galactosidase from cell extract using hollow fiber ion‐exchange membrane adsorption
Authors: Heng Meng H. , Glatz Charles E.
Published: December 2004

We explored the use of charged fusions for selective recovery of β‐galactosidase from cell extract using a low‐cost, easily scaled, fast, charge‐based separation technique—ion exchange on hollow fiber ion‐exchange membranes (HFIEMs). The additional charges carried by a series of anionic fusion tails allowed selective binding and release of β‐galactosidase from Escherichia coli cell extract using the HFIEM cartridge. The purification factors increased with fusion length. The β‐galactosidase was recovered in active form. For the longest fusion studied, more than sixfold enrichment in specific activity was attained. The specific activity of the recovered fraction is comparable with that of commercial wild‐type β‐galactosidase and affinity‐purified fusion protein. © 1993 John Wiley & Sons, Inc.
The molecular weight cut‐off of microcapsules is determined by the reaction between alginate and polylysine
Authors: Vandenbossche Geert M. R. , Van Oostveldt Patric , Demeester Joseph , Remon Jean‐Paul
Published: December 2004

Mammalian cells encapsulated in alginate‐polylysine microcapsules are used as artificial organs in cancer research and in biotechnology. These applications require microcapsules with a reproducible mol. wt. cut‐off. The high cost of the polycation, polylysine, requires an efficient preparation procedure. This article shows that the overall reported contact time of 5 minutes at ambient conditions should be increased several times in order to reach a maximal binding between the calcium alginate beads and 0.1% (w/v) polylysine solutions. An increase of the polylysine concentration from 0.0125% to 0.8% (w/v) resulted in a faster maximal binding, but the amount of polylysine bound increased also. Immersion of calcium alginate beads with a diameter of 750 μm, prepared from 1 mL alginate, in 30 mL of a 0.8% (w/v) polylysine solution, resulted in a polylysine spill of more than 89%. The time required to reach a maximal binding was related to the reaction temperature. The interaction zone between calcium alginate beads and fluorescein isothiocyanate‐labeled polylysine solutions was visualized with a confocal laser scanning microscope as a function of time. Microcapsules, prepared at 40°C with 0.1% (w/v) polylysine solutions with mol. wts. between 12 and 249.2 kD, were permeable for fluorescein isothiocyanate‐labeled dextran, mol. wt. 4.7, but not for 40.5 kD. Higher polylysine concentrations resulted in a membrane with a mol. wt. cut‐off lower than 4.7 kD. © 1993 John Wiley & Sons, Inc.
Immobilization of enzyme onto cellulose–titanium oxide composite fiber
Authors: Kurokawa Y. , Sano T. , Ohta H. , Nakagawa Y.
Published: December 2004

Fibers of a cellulose–TiO2 composite were prepared by the reaction of cellulose with titanium iso‐propoxide. Enzymes were immobilized on the fibers easily and simply under mild conditions. The fibers were stable in common solvents, high ionic solutions, and over a wide range of pH values 3–10. © 1993 John Wiley & Sons, Inc.
Generation of a pH gradient in an immobilized enzyme system
Authors: Byers James P. , Shah Manish B. , Fournier Ronald L. , Varanasi Sasidhar
Published: December 2004

Several examples of two‐step sequential reactions exist where, because of the poor equilibrium conversion by the first reaction, it is desirable to conduct the two reactions simultaneously. In such a scheme, the product of the first reaction is continuously removed by the second reaction, thus not allowing the first reaction to approach chemical equilibrium. Therefore, the first reaction is allowed to proceed in the desired direction at an appreciable rate. However, in many biochemical applications where enzyme catalysts are involved, the enzyme's activities are strong functions of pH. Where the pH optima of the first and second reaction differ by three to four units, the above reaction scheme would be difficult to implement. In these cases, the two reactions can be separated by a thin permeable membrane across which the desired pH gradient is maintained. In this article, it was shown, both by theory and experiment, that a thin, flat membrane of immobilized urease can accomplish this goal when one face of the membrane is exposed to the acidic bulk solution (pHb = 4.5) containing a small quantity of urea (0.01 M). In this particular case, the ammonia that was produced in the membrane consumed the incoming hydrogen ions and thus maintained the desired pH gradient. Experimental results indicate that with sufficient urease loading, the face of the membrane opposite to the bulk solution could be maintained at a pH that would allow many enzymes to realize their maximum activities (≈ 7.5). It was also found that this pH gradient could be maintained even in the presence of a buffer, which greatly enhances the transport of protons into the membrane. © 1993 John Wiley & Sons, Inc.
Detection of bacterial contamination in cultures of eucaryotic cells by gas chromatography–mass spectrometry
Authors: Elmroth Ingrid , Fox Alvin , Holst Olle , Larsson Lennart
Published: December 2004

The use of gas chromatography–mass spectrometry for early detection of bacterial contaminations in cultures of baker's yeast, Penicillium chrysogenum, and an animal cell line was evaluated; muramic acid and characteristic cellular fatty acids were used as analytes. By analyzing branched‐chain and cyclopropane‐substituted fatty acids as methyl esters, Staphylococcus epidermidis, Bacillus subtilis, Lactobacillus reuteri, Enterobacter cloacae, and Pseudomonas fluorescens were detected in a 500‐fold excess (w/w) of baker's yeast; the amounts injected corresponded to 300 ng (dry mass) of the bacteria. Contamination with Bacillus was detected in cultures of Penicillium chrysogenum and animal cells by analyzing muramic acid, both as its alditol acetate derivative, using electron impact ionization, and its trifluoroacetyl methyl glycoside derivative, using negative ion‐chemical ionization. The trifluoroacetylated derivative was detected in injected amounts corresponding to 1 × 103 bacterial cells in the contaminated animal cell line, whereas amounts corresponding to 1 × 105 bacteria were required for detection of the alditol acetate derivative; the amounts in the original samples were 5 × 105 and 5 × 106, respectively. However, the alditol acetate method exhibited lower chemical interferences than the trifluoroacetyl methyl glycoside procedure. The results show the potential of using gas chromatographic–mass spectrometric analysis of cellular constituents for the detection of bacterial contaminations in eucaryotic cultures as an alternative to conventional microbiological methods. © 1993 John Wiley & Sons, Inc.
Microbial utilization of levoglucosan in wood pyrolysate as a carbon and energy source
Authors: Prosen Elizabeth M. , Radlein Desmond , Piskorz Jan , Scott Donald S. , Legge Raymond L.
Published: December 2004

The Waterloo Fast Pyrolysis Process (WFPP) can produce an organic liquid high in levoglucosan (1, 6‐anhydro‐β‐D‐glucopyranose) content from suitably pretreated lignocellulosics. A variety of fungi and yeasts were screened for their ability to utilize and ferment this organic liquid. To enhance its fermentability, the pyrolysis tar was posttreated in three different ways: (1) an aqueous extract (lignin removed); (2) activated charcoal treated (lignin and aromatics removed); and (3) acid hydrolysate (lignin and aromatics removed with the levoglucosan hydrolyzed to glucose). Four fungal strains were examined. None grew in the aqueous extract, but all grew equally well in both the activated charcoal treated and the acid hydrolysate, suggesting that the aromatic species were inhibitory to growth. Seven yeast species were examined, two of which did not grow on any of the extracts. Five of the yeast strains grew well on both the aqueous extract as well as the activated charcoal extract. The hydrolysate was optimal in terms of biomass yield and ethanol production. Ethanol yields on the hydrolysate were comparable or better than those on glucose. Ethanol was also produced in the aqueous extract and activated charcoal‐treated substrate, but yields were considerably lower than on the hydrolysate or glucose. It is apparent that a wood pyrolysate maximized for levoglucosan can serve as a fermentable substrate, although postpyrolysis clean‐up appears necessary. © 1993 John Wiley & Sons, Inc.
A fractal model for the characterization of mycelial morphology
Authors: Patankar Dhananjay B. , Liu Tuan‐Chi , Oolman Timothy
Published: December 2004

A new technique based on a fractal model has been developed for the quantification of the macroscopic morophology of mycelia. The morphological structuring is treated as a fractal object, and the fractal dimension, determined by an ultrasonic scattering procedure developed for the purpose, serves as a quantitative morphological index. Experimental observations reported earlier and simulations of mycelial growth, carried out using a probabilistic‐geometric growth model developed for the purpose, both validate the applicability of the fractal model. In experiments with three different species, the fractal dimensions of pelletous structures were found to be in the range 1.45–2.0 and those of filamentous structures were in the range 1.9–2.7, with values around 2.0 representing mixed morphologies. Fractal dimensions calculated from simulated mycelia are in rough agreement with these ranges. The fractal dimension is also found to be relatively insensitive to the biomass concentration, as seen by dilution of the original broths. The relation between morphology and filtration properties of the broths has also been studied. The fractal dimension shows a strong correlation with the index of cake compressibility and with the Kozeny constant, two filtration parameters that are known to be morphology dependent. This technique could thus be used to develop correlations between the morphology, represented by the fractal dimension, and important morphology‐dependent process variables. © 1993 John Wiley & Sons, Inc.
Partitioning behavior and enrichment of proteins with reversed micellar extraction: I. Forward extraction of proteins from aqueous to reversed micellar phase
Authors: Nishiki Tadaaki , Sato Ikuto , Kataoka Takeshi , Kato Daisaburo
Published: December 2004

Protein extractions using aerosol OT (AOT)–isooctane reverse micelle solutions have been studied to explore the potential for separating and enriching proteins with the reversed micellar extraction. The effects of pH, ionic strength, and different cations of chlorides in a bulk aqueous phase and of AOT concentration in an organic phase on the partitioning of lysozyme and myoglobin and the solubilization of water are presented in detail. The extraction of lysozyme was affected by the concentration of potassium or barium but was almost independent of that of sodium or calcium, whose ionic diameter is smaller than that of potassium and barium. For the extraction of myoglobin, however, the effect of barium concentration was not appreciable. Lysozyme could be enriched into the reversed micellar phase up to 30 times the aqueous feed concentration. © 1993 John Wiley & Sons, Inc.
Effect of surfactants on cellulose hydrolysis
Authors: Helle Steve S. , Duff Sheldon J. B. , Cooper David G.
Published: December 2004

The effect of surfactants on the heterogeneous enzymatic hydrolysis of Sigmacell 100 cellulose and of steam‐exploded wood was studied. Certain biosurfactants (sophorolipid, rhamnolipid, bacitracin) and Tween 80 increased the rate of hydrolysis of Sigmacell 100, as measured by the amount of reducing sugar produced, by as much as seven times. The hydrolysis of steam‐exploded wood was increased by 67% in the presence of sophorolipid. At the same time, sophorolipid was found to decrease the amount of enzyme adsorbed onto the cellulose at equilibrium. Sophorolipid had the greatest effect on cellulose hydrolysis when it was present from the beginning of the experiment and when the enzyme/cellulose ratio was low. © 1993 John Wiley & Sons, Inc.
Optimization of alkyl β‐D‐galactopyranoside synthesis from lactose using commercially available β‐galactosidases
Authors: Stevenson David E. , Stanley Roger A. , Furneaux Richard H.
Published: December 2004

Commercially available lactase (β‐D‐galactoside galactohydrolase, EC 3.2.1.23) enzymes produced from Kluyveromyces fragilis and Kluyveromyces lactis were accessed as catalysts for use in the production of β‐galactopyranosides of various alcohols using lactose as galactosyl donor. The yield of galactoside was enhanced by using the highest practical concentrations of both lactose and alcohol acceptor. The concentrations and thus yield, were limited by the solubility of the substrates. The increase in galactoside yield with increasing lactose concentration appeared to be specific to the lactose substrate and not due to water activity alterations, because addition of maltose to a fixed concentration of lactose had no effect. During the course of the reaction, the yield of galactoside peaked after around 70% to 80% of the lactose was consumed, due to hydrolysis of the product by the enzyme. A wide variety of compounds with primary or secondary hydroxyl groups could act as acceptors, the essential requirement being at least some water solubility. Addition of organic cosolvents had little effect on galactoside yield except when it increased the water solubility of sparingly soluble alcohols. Some galactosides were synthesized on a gram scale to determine practical product recoveries and improve purification methods for large‐scale synthesis. Initial purification by hydrophobic chromatography (for galactosides of hydrophobic alcohols) or strong anion‐exchange chromatography (for galactosides of hydrophilic alcohols) separated galactosides, galactobiosides, and higher oligomers from reducing sugars. A facile separation of the galactoside and galactobioside could then be effected by flash chromatography on silica gel. © 1993 John Wiley & Sons, Inc.
Toxicity of homologous series of organic solvents for the gram‐positive bacteria Arthrobacter and Nocardia Sp. and the gram‐negative bacteria Acinetobacter and Pseudomonas Sp.
Authors: Vermuë Marian , Sikkema Jan , Verheul Annette , Bakker Rudolf , Tramper Johannes
Published: December 2004

The toxicity of homologous series of organic solvents has been investigated for the gram‐positive bacteria, Arthrobacter sp. and Nocardia sp., and the gram‐negative bacteria, Acinetobacter sp. and Pseudomonas sp. The hydrophobicity of the solvent, expressed by its logPoctanol, proves to be a good measure for the toxicity of solvents in a two‐phase system. The transition from toxic to nontoxic solvents occurs between logPoctanol 3 and 5 and depends on the homologous series. No correlation has been found between the hydrophobicity of the substituent on the alkyl backbone of the solvent and the location of the transition point in toxicity. The logPoctanol, above which all solvents are nontoxic, is used to express the solvent tolerance of the bacteria. In general, the solvent tolerance of gram‐negative bacteria is found to be slightly higher than that of gram‐positive bacteria, but this does not hold for all homologous series of organic solvents investigated.

Because the toxicity effects of organic solvents in a two‐phase system can be ascribed to molecular as well as phase toxicity effects, molecular toxicity effects were investigated separately in a one‐phase system with subsaturating amounts of organic solvent. The solvent concentration in the aqueous phase, at which 50% of the metabolic activity of the bacteria is lost, is used to express solvent toxicity. This concentration is found to be similar for the gram‐positive Arthrobacter and the gram‐negative Acinetobacter. Assuming the critical membrane concentration theory (G. J. Osborne et al. Enzyme Microb. Technol. 1990, 12: 281–291) to be valid, it can be concluded that differences in solvent tolerance between these two bacteria, cannot be ascribed to differences in response to molecular toxicity. Prediction of the toxicity of any solvent, using the critical membrane theory, appears to be possible in the case of alkanols or alkyl acetates. However, prediction of the toxicity of ethers appears to be impossible. © 1993 John Wiley & Sons, Inc.
Morphological measurements on Penicillium chrysogenum broths by rheology and filtration methods
Authors: Liu Tuanchi , Yu Duncan
Published: December 2004

The morphology parameters of mycelial culture (Penicillium chrysogenum) were measured and quantified by rheology and filtration methods. Two of the morphology parameters obtained from rheology measurements, δ defined by the Casson equation and δ* defined by intrinsic viscosity, were found to vary systematically with broth age and with the observed morphology by microscopy. Three of the filtration parameters, hyphal density, Kozeny constant, and index of compressibility, are demonstrated as sensitive indicators of the broth age and mycelial morphology. Two of the morphology parameters, δ and δ*, were used to cross‐correlate with hyphal density. Because various mycelial fermentations require different growth morphologies (pellet and filament) for optimum product yield and the morphology of mycelial broths varies with broth age, it is suggested that these morphology parameters could be used to represent the morphology of mycelial broths quantitatively. © 1993 John Wiley & Sons, Inc.
Efficiency of fatty acid synthesis by oleaginous yeasts: Prediction of yield and fatty acid cell content from consumed C/N ratio by a simple method
Authors: Granger L‐M. , Perlot P. , Goma G. , Pareilleux A.
Published: December 2004

In nitrogen‐limited media, growth and fatty acid formation by the oleaginous yeast Rhodotorula glutinis, i.e., yield and fatty acid cell content, have been characterized regarding carbon and nitrogen availabilities. It was shown that the formation of fatty acid free biomass was limited by nitrogen availability, whereas the fatty acid production was directly dependent on the consumed C/N ratio. According to these observations, the fraction of substrate consumed for fatty acid synthesis was estimated by using a simple method based on the actual yields, i.e., the mass of carbon source strictly converted into fatty acids and fatty acid free biomass. From these results, relationships were established allowing to predict in a simple and performing manner the maximal attainable fatty acid cell content and yield from the available carbon and nitrogen. These relationships were validated by using experimental data obtained by various authors with different yeast strains, and the proposed method was compared to the energetic and mass balance method previously described. © 1993 John Wiley & Sons, Inc.
Affinity–based reversed micellar protein extraction: I. Principles and protein–ligand systems
Authors: Kelley Brian D. , Wang Daniel I. C. , Hatton T. Alan
Published: December 2004

Affinity cosurfactants, consisting of hydrophilic ligands derivatized with hydrophobic tails, increase the efficiency of selective protein recoveries using reversed micelles by extending the operating range of pH and salt concentration over which an extraction can be performed. Three different affinity cosurfactant–protein pairs have been used to demonstrate the principles of this extractive technique: (i) concanavalin A, a lectin, was extracted with the addition of octyl glucoside; (ii) natural amphiphiles, such as lecithin, were used to extract myelin basic protein, a membrane‐associated protein known to recognize and bind the phosphatidylcholine headgroup; and (iii) alkyl boronic acids were used to extract chymotrypsin. The enhancement in protein transfer correlated with the binding strength of the free ligand and protein in aqueous solution. Several control studies confirmed the biospecificity of the interactions of protein and affinity cosurfactant. © 1993 John Wiley & Sons, Inc.
Morphological characterization of yeast by image analysis
Authors: Pons M. N. , Vivier H. , Rémy J. F. , Dodds J. A.
Published: December 2004

A semiautomatic image analysis method, with minimal operator intervention, has been developed to characterize the morphology of yeast cells under the assumption that they have an ellipsoidic shape. The cells are observed by optical microscopy and the surface and the minor and major half‐axes of the projection of the ellipsoid on the image plane are determined. Using this method, yeast size distributions and population kinetics (single and budding cells, cell clusters) are determined during alcoholic fermentations. Combination of image analysis with a methylene blue viability test is examined but the staining procedure induces a change in the size of the cells. © 1993 John Wiley & Sons, Inc.
Effect of solution environment on the permeability of red blood cells
Authors: Langsdorf Leah J. , Zydney Andrew L.
Published: December 2004

The cell membrane permeability governs the rate of solute transport into and out of the cell, significantly affecting the cell's metabolic processes, viability, and potential usefulness in both biotechnological applications and physiological systems. Most previous studies of the cell membrane permeability have neglected the possible effects of suspending medium on membrane transport, even though there is extensive experimental evidence that suspending phase composition can significantly affect other properties related to the cell membrane (e.g., cell deformability, fragility, and aggregation rate). This study examined the effects of suspending phase composition (both proteins and electrolytes) on the permeability of human red blood cells to the metabolites creatinine and uric acid. Data were obtained using a stirred ultrafiltration device with direct cell‐ and proteinfree sampling through a semipermeable membrane. Both the uric acid and creatinine permeabilities were strongly affected by the suspending phase composition, with the permeabilities in different buffer solutions varying by as much as a factor of three. The predominant factors affecting the permeability were the presence (or absence) of chloride, phosphate/adenine, and proteins, although the magnitude and even the direction of these effects were significantly different for creatinine and uric acid transport. The dramatic differences in behavior for uric acid and creatinine reflect the different transport mechanisms for these solutes, with uric acid transported by a carrier‐mediated mechanism and creatinine transported by passive diffusion through the lipid bilayer. These results provide important insights into the effects of solution environment on cell membrane transport and other cell membrane‐mediated properties. © 1994 John Wiley & Sons, Inc.
Mass transfer effects in solvent‐free fat interesterification reactions: Influences on catalyst design
Authors: Ison A. P. , Macrae A. R. , Smith C. G. , Bosley J.
Published: December 2004

The use of solvent‐free systems in the oil and fats industry is commonplace. Initial studies on interesterification were carried out in solvent systems because the lipase was immobilized solely by adsorption onto particles of diatomaceous earth. In this study, the mass transfer characteristics associated with the continuous interesterification of olive oil in a solvent‐free system have been examined, for lipase immobilized on the three ion‐exchange materials: Duolite ES562, Duolite ES568, and Spheroil DEA. The process of immobilization is influenced by the internal structure of the material and this in turn influences the interesterification activity of the catalyst. Individually prepared catalysts for the three support materials have shown that external mass transfer limitations are unlikely even at low flowrates.

In the case of Spherosil DEA, with a mean pore diameter of 1480 Å, the wide pores would be expected to reduce internal mass transfer limitations; however, it is more likely that the reduction in activity with increased catalyst loading is due to the lipase molecules being immobilized in a tightly packed monolayer. In such a situation, some active sites of the lipase molecules would become inaccessible to substrate molecules leading to an observed reduction in activity. For Duolite ES568, the observed results are very similar to those seen for Spherosil DEA, however, the pore structure of this support material indicate that some internal mass transfer limitations may also be occurring. Yet the contribution of the individual effects cannot be determined. The results observed for the support Duolite ES562 are different than those observed for the other materials and reflect the heterogeneity of Duolite ES562. The large proportion of narrow pores in the support mean that, for the catalysts examined, immobilization is most likely to have occurred in the external pores of the particles, and as such no internal mass transfer limitation is observed.

It is clear that for interesterification the material chosen for enzyme immobilization will have an important role in determining the catalyst efficiency. External mass transfer limitations are very minor and observed internal mass transfer limitations may be caused by both internal mass transfer and the manner in which the immobilization process occurs. © 1994 John Wiley & Sons, Inc.
Oxygen uptake and citric acid production by Candida lipolytica Y 1095
Authors: Rane Kishore D. , Sims Kevin A.
Published: December 2004

The rates of oxygen uptake and oxygen transfer during cell growth and citric acid production by Candida lipolytica Y 1095 were determined. The maximum cell growth rate, 1.43 g cell/L · h, and volumetric oxygen uptake rate, 343 mg O2/L · h, occurred approximately 21 to 22 h after inoculation. At the time of maximum oxygen uptake, the biomass concentration was 1.3% w/v and the specific oxygen uptake rate was slightly greater than 26 mg O2/g cell · h. The specific oxygen uptake rate decreased to approximately 3 mg O2/g cell · h by the end of the growth phase.

During citric acid production, as the concentration of dissolved oxygen was increased from 20% to 80% saturation, the specific oxygen uptake and specific citric acid productivity (mg citric acid/g cell · h) increased by 160% and 71%, respectively, at a biomass concentration of 3% w/v. At a biomass concentration of 5% w/v, the specific oxygen uptake and specific citric acid productivity increased by 230% and 82%, respectively, over the same range of dissolved oxygen concentrations.

The effect of dissolved oxygen on citric acid yields and productivities was also determined. Citric acid yields appeared to be independent of dissolved oxygen concentration during the initial production phase; however, volumetric productivity (g citric acid/L · h) increased sharply with an increase in dissolved oxygen. During the second or subsequent production phase, citric acid yields increased by approximately 50%, but productivities decreased by roughly the same percentage due to a loss of cell viability under prolonged nitrogen‐deficient conditions. © 1994 John Wiley & Sons, Inc.
Ethanol production from nonsterilized carob pod extract by free and immobilized Saccharomyces cerevisiae cells using fed‐batch culture
Authors: Roukas T.
Published: December 2004

The production of ethanol from carob pod extract by free and immobilized Saccharomyces cerevisiae cells in batch and fed‐batch culture was investigated. Fed‐batch culture proved to be a better fermentation system for the production of ethanol than batch culture. In fed‐batch culture, both free and immobilized S. cerevisiae cells gave the same maximum concentration (62 g/L) of final ethanol at an initial sugar concentration of 300 g/L and F = 167 mL/h. The maximum ethanol productivity (4.4 g/L h) was obtained with both free and immobilized cells at a substrate concentration of 300 g/L and F = 334 mL/h. In repeated fed‐batch culture, immobilized S. cerevisiae cells gave a higher overall ethanol concentration compared with the free cells. The immobilized S. cerevisiae cells in Ca‐alginate beads retained their ability to produce ethanol for 10 days. © 1994 John Wiley & Sons, Inc.
Selective protein transport through a thin cellulose coated porous membrane
Authors: Yang Yiting , Jacobs Harvey , Bae You Han , Kim Sung Wan
Published: December 2004

A new composite membrane was designed and studied for permselectivity of various molecular weight proteins. The membrane is composed of a porous substrate membrane [Durapore; poly(vinylidene fluoride)] coated with a thin dense layer of regenerated cellulose. This composite membrane was fabricated by spin coating a cellulose acetate solution onto the membrane, followed by alkaline hydrolysis of the cellulose acetate coating to regenerate cellulose. The coated layer was physically characterized by scanning electron microscopy (SEM) and infrared (IR) spectroscopy. In addition, the water uptake into and permeation properties of macromolecules across the coated and uncoated membranes were studied. A typical composite membrane coating was 0.8 ± 0.2 μm thick, resulting in a molecular weight cutoff of approximately 40,000 daltons. This composite membrane also demonstrated negligible diffusional lag time for permeants, due to the diffusional barrier. © 1994 John Wiley & Sons, Inc.
Biosensor for continuos glucose monitoring
Authors: Atanasov Plamen , Wilkins Ebtisam
Published: December 2004

A potentially implantable glucose biosensor for continuous monitoring of glucose levels in diabetic patients has been developed. The glucose biosensor is based on an amperometric oxygen electrode and glucose oxidase immobilized on carbon powder held in a form of a liquid suspension. The enzyme material can be replaced (the sensor recharged) without sensor disassembly. Recharging of the biosensor is achieved by injecting fresh immobilized enzyme into the sensor using a septum. Diffusion membranes made of silastic latex‐rubber coatings over a microporous polycarbonate membrane are used. Calibration curves of the amperometric signal show linearity over a wide range of glucose concentrations—up to 500 mg/dL (28 mM), covering hypoglycemic, normoglycemic, and hyperglycemic conditions. Preliminary in vitro studies of the biosensor show stable performance during several recharge cycles (of 14 days each) over a period of 4 months. © 1994 John Wiley & Sons, Inc.
Modeling hexavalent chromium reduction in Escherichia coli 33456
Authors: Shen Hai , Wang Yi‐Tin
Published: December 2004

A model based on te analysis of the mechanism of enzymatic reactions was developed to characterize the rate and extent of microbial reduction of hexavalent chromium in Escherichia coli 33456. A finite reduction capacity (Rc) was proposed and incorporated into the enzymatic model to regulate the toxicity effect on cells due to the oxidizing power of Cr(VI). The parameter values were determined by nonlinear least‐square analysis using experimental data of anaerobic cultures. The obtained parameters were then used to predict Cr(VI) reduction in aerobic cultures along with a modification term of uncompetitive inhibition from molecular oxygen. The applicability of the developed model was demonstrated through excellent prediction of the results of batch studies conducted over range of initial Cr(VI) concentrations, initial cell densities, and DO levels. A sensitivity analysis revealed that the parameters obtained using the experimental data were unique, and neither change in Kc, the half‐velocity constant, at high initial Cr(VI) concentrations nor change in Rc, the reduction capacity, at low initial Cr(VI) concentrations was sensitive to model prediction. © 1994 John Wiley & Sons, Inc.
Optimization of an industrial microalgae fermentation
Authors: Hilaly A. K. , Karim M. N. , Guyre D.
Published: December 2004

Optimization of cellular productivity of an industrial microalgae fermentation was investigated. The fermentation was carried out at Coors Biotech Products Company, Fort Collins, Colorado. A mathematical model was developed based on the data collected from pilot plant test runs at different operating conditions. Pontryagin's maximum principle was used for determining the optimal feed policy. A feedback control algorithm was also studied for maximizing the cellular productivity. During continuous operation, the optimum dilution rate was determined by an adaptive optimization scheme based on the steepest descent technique and a recursive least squares estimation of model parameters. A direct search algorithm was also applied to determine the optimum feed rate. Comparison of the theoretical results of the different optimization schemes revealed that the direct search algorithm was preferable because of its simplicity. The experimental results of real time application of the feedback algorithm agreed fairly well with those of the theoretical analyses. © 1994 John Wiley & Sons, Inc.
Inactivation of enzymes by organic solvents: New technique with well‐defined interfacial area
Authors: Ghatorae Amreek S. , Bell George , Halling Peter J.
Published: December 2004

A liquid‐liquid bubble column apparatus allows exposure of enzyme solutions to water‐immiscible organic solvents with a known total interfacial area and welldefined time scales and flow. It allows clear distinction of the different classes of inactivation mechanism. With urease as a model enzyme, octan‐2‐one and butylbenzene act only through the effects of solvent molecules dissolved in the aqueous phase, giving first‐order inactivation at 0.34 and 0.21 h−1, respectively. Hexane and tridecane act only through exposure to the interface. The amount of urease inactivated is proportional to the total area of interface exposed, rather than to elapsed time, and may be characterized by a rate of about 0.5 μkat m−2. This is consistent with the formation and (partial) inactivation of a complete adsorbed monolayer of protein. With butan‐1‐ol, both mechanisms contribute significantly to the observed inactivation. The presence of O2 increases the rate of interfacial inactivation, but not that by dissolved solvent. © 1994 John Wiley & Sons, Inc.
Electrochemical disinfection of bacteria in drinking water using activated carbon fibers
Authors: Matsunaga Tadashi , Nakasono Satoshi , Kitajima Yoji , Horiguchi Kazuo
Published: December 2004

A novel electrochemical reactor employing activated carbon fiber (ACF) electrodes was constructed for disinfecting bacteria in drinking water. Escherichia coli adsorbed preferentially onto ACF rather than to carbon‐cloth or granular‐activated carbon. E. coli cells, which adsorbed onto the ACF, were killed electrochemically when a potential of 0.8 V vs. a saturated calomel electrode (SCE) was applied. Drinking water was passed through the reactor in stop‐flow mode: 2mL/min for 12 h, o L/min for 24 h, and 1 mL/min for 6 h. At an applied potential of 0.8 V vs, SCE, viable cell concentration reamined below 30 cells/mL. In the absence of an applied potential, bacteria grew to a maximum concentration of 9.5 × 103 cells/mL. After continuous operation at 0.8 V vs. SCE, cells adsorbed onto the ACF could not be observed by scanning electron microscopy. In addition, chlorine in drinking water was completely removed by the reactor. Therefore, clean and efficient inactivation of bacteria in drinking water was successfully performed. © 1994 John Wiley & Sons, Inc.
Monitoring recombinant inclusion body recovery in an industrial disc stack centrifuge
Authors: Jin K. , Thomas O. R. T. , Dunnill P.
Published: December 2004

A simple and rapid spectrophotometric method for measuring recombinant inclusion body concentrations in the presence of Escherichia coli cell debris has been applied to monitoring the performance of an industrial disc stack centrifuge. Turbidimetric measurements were made at two wavelengths, i.e., 600 nm and 420 nm, and the ratios of OD600nm/OD420nm related to the particle composition in suspension. The principle behind the technique is that inclusion body particles scatter light at 600 nm more effectively than do smaller cell debris particles when compared with the degree of light scatter at 420 nm. This technique may have broad potential application in developing an automatic monitoring and control system for industrial‐scale inclusion body recovery. © 1994 John Wiley & Sons, Inc.
Biological destruction of CCl4: I. Experimental design and data
Authors: Petersen James N. , Skeen Rodney S. , Amos Kristine M. , Hooker B. S.
Published: December 2004

A denitrifying consortium capable of transforming carbon tetrachloride (CCl4) was cultured from aquifer sediment from the U.S. Department of Energy's Hanford Site in southeastern Washington State. To understand the kinetics of the biological destruction of CCl4 by these microbes, a set of experiments, the conditions of which were chosen according to a fractional factorial experimental design, were completed. This article reports on the experimental design along with the results for CCl4, biomass, acetate, nitrate, and nitrite concentrations. These data indicate that growth is inhibited by high nitrite concentrations, whereas CCl4 degradation is slowed by the presence of nitrate and/or nitrite. © 1994 John Wiley & Sons, Inc.
Mechanism of extraction of chymotrypsin into isooctane at very low concentrations of aerosol OT in the absence of reversed micelles
Authors: Paradkar Vikram M. , Dordick Jonathan S.
Published: December 2004

Chymotrypsin is easily extracted from an aqueous solution into isooctane containing the anionic surfactant aerosol OT (AOT). The concentration of AOT needed to efficiently extract 0.5 mg/mL CMT is as low as 1 mM and as low as 0.2 mM AOT was sufficient to extract the protein into isooctane. The extraction process was unaffected by 10% (v/v) ethyl acetate in the isooctane phase. Moreover, spectroscopic analysis by electron paramagnetic resonance indicated that CMT did not exist inside a discreet water pool of a reversed micelle. Calculations of the number of AOT molecules associated per extracted CMT molecule indicate that only ca. 30 surfactant molecules interact with the protein, a value too low for reversed micellar incorporation of the protein in isooctane. These studies suggested that reversed micelles do not need to be involved in the actual transfer of the protein from the aqueous to the organic phase and protein solubilization in the organic phase is possible in the absence of reversed micelles. Based on these findings, a new mechanism has been proposed herein for protein extraction via the phase transfer method involving ionic surfactants. The central theme of this mechanism is the formation of an electrostatic complex between CMT and AOT at the aqueous/organic interface between AOT and CMT, thereby leading to the formation of a hydrophobic species that partitions into the organic phase. Consistent with this mechanism, the efficiency of extraction is dependent on the interfacial mass transfer, the concentrations of CMT and AOT in the aqueous and organic phases, respectively; the ionic strength of the aqueous phase; and the presence of various cosolvents. © 1994 John Wiley & Sons, Inc.
Serum‐free media for cultures of primitive and mature hematopoietic cells
Authors: Sandstrom Craig E. , Miller William M. , Papoutsakis Eleftherios T.
Published: December 2004

The in vitro culture of human hematopoietic cells has many research and therapeutic applications. Traditionally, human hematopoietic cultures have been conducted using serum‐containing media. The disadvantages inherent in the use of serum could be eliminated by the use of serum‐free media. In this review, we summarize and discuss the current status of serum‐free media for both mature and immature human hematopoietic cells. The mature hematopoietic cells discussed are of lymphoid (e.g., lymphokine activated killer cells and tumor infiltrating lymphocytes) and myeloid origin (e.g., monocytes/macrophages). The cultures of immature hematopoietic cells discussed are clonogenic and long‐term cultures. In addition, we briefly review the types of human hematopoietic cells, their clinical applications, and the basic strategies and components used to formulate serum‐free media, Finally, we outline future requirements and directions in the development of serum‐free media for primitive hematopoietic cells.
Development of a bilayered living skin construct for clinical applications
Authors: Wilkins Leon M. , Watson Stephen R. , Prosky Stefan J. , Meunier Susan F. , Parenteau Nancy L.
Published: December 2004

An in vitro construct of human skin (living skin equivalent, LSE) has been engineered using serially passaged human epidermal keratinocytes and human dermal fibroblasts with a matrix of type I collagen. Cells are obtained from neonatal foreskin. LSE is cast, cultured, and shipped in a single culture insert. The size and shape of theinsert determines thesize and shape of the LSE. The dermal matrix consists of dermal fibroblasts within a condensed collagen lattice. The overlying epidermis is developed at the air–liquid interface to generate a protective cornified layer. Serum was not necessaryfor development of the epidermis. LSE for graft (Graftskin) has handling characteristics similar to split‐thickness skin allowing it to be meshed, stapled, and sutured. LSE was cryopreserved using 65% glycerol an rapid freezing. Viability and in vivo performance on athymic mice were similar to fresh LSE. Cells derived from human eccrine gland were able to invade and form tubules rudimentary appendages may be possible.
Leuconostoc mesenteroides growth kinetics with application to bacterial profile modification
Authors: Lappan Raymond E. , Fogler H. Scott
Published: December 2004

Bacterial profile modification (BPM) is being developed as an oil recovery technique that uses bacteria to selectively plug oil depleted zones within a reservoir to divert displacing fluids (typically water) into oil‐rich zones. Leuconostoc mesenteroides, which produces dextran when supplied with sucrose, is a bacterium that is technically feasible for use in profile modification. However, the technique requires controlled bacterial growth to produce selective plugging.

A kinetic model for the production of cells and polysaccharides has been developed for L. mesenteroides bacteria. This model, based on data from batch growth experiments, predicts saccharide utilization, cell generation, and dextran production. The underlying mechanism is the extracellular breakdown of sucrose into glucose and fructose and the subsequent production of polysaccharide (dextran). The monosaccharides are then available for growth. Accompanying sucrose consumption is the utilization of yeast extract. The cell requires a complex media that is provided by yeast extract as a source of vitamins and amino acids. Varying the concentration ratio of yeast extract to sucrose in the growth media provides a means of controlling the amount of polymer produced per cell. Consequently, in situ bacteria growth can be controlled by the manipulation of nutrient media composition, thereby providing the ability to create an overall strategy for the use of L. mesenteroides bacteria for profile modification.
Production of poly(3‐hydroxybutyric acid) by fed‐batch culture of Alcaligenes eutrophus with glucose concentration control
Authors: Kim Beom Soo , Lee Seung Chul , Lee Sang Yup , Chang Ho Nam , Chang Yong Keun , Woo Seong Ihl
Published: December 2004

Alcaligenes eutrophus NCIMB 11599 was cultivated to produce poly(3‐hydroxybutyric acid) (PHB) from glucose by the automatic fed‐batch culture technique. The glucose concentration of the culture broth was controlled at 10 to 20 g/L by two methods: using exit gas data obtained from a mass spectrometer and using an on‐line glucose analyzer. The effect of ammonium limitation on PHB synthesis at different culture phases was studied. The final cell concentration, PHB concentration, and PHB productivity increased as ammonia feeding was stopped at a higher cell concentration. High concentrations of PHB (121 g/L) and total cells (164 g/L) were obtained in 50 h when ammonia feeding was stopped at the cell concentration of 70 g/L. The maximum PHB content reached 76% of dry cell weight and the productivity was 2.42 g/L h with the yield of 0.3 g PHB/g glucose.
Protease‐catalyzed synthesis of oligopeptides in heterogenous substrate mixtures
Authors: López‐Fandiño Rosina , Gill Iqbal , Vulfson Evgeny N.
Published: December 2004

A systematic study of enzymatic peptide synthesis in heterogeneous substrate mixtures was carried out, with the aim of establishing the preparative scope of this methodology. Semiliquid eutectics were obtained with various combinations of neutral, acidic, and basic amino acid derivatives, in the presence or absence of adjuvants. A range of serine cysteine, and metalloproteases readily catalyzed the formation of the required dipeptides under these conditions. The synthetic usefulness of the approach was demonstrated by the sequential and convergent synthesis of derivatives of a number of bioactive di‐, tri‐, and pentapeptides, including aspartame, sweet lysine peptide, kyotorphin amide, ACE‐inhibiting and ‐immunoactive tripeptides, and Leu‐enkephalin amide, with overallyields of 21% to 84% and productivities of 0.13 to 0.75 g/g being obtained. © 1994 John Wiley & Sons, Inc.