bluuebunny/crossref_metadata_2025 huggingface dataset:
Micropatterning Proteins and Synthetic Peptides on Solid Supports: A Novel Application for Microelectronics Fabrication Technology
Authors: Britland Stephen , Perez‐Arnaud Enrique , Clark Peter , McGinn Brian , Connolly Patricia , Moores Geoffrey
Published: March 2005

In this paper, we describe a method for immobilizing proteins and synthesizing peptides in micrometer‐dimension patterns on solid supports. Microelectronics fabrication technology was adapted and used to lithographically direct the location of immobilization of proteins on appropriately derivatized surfaces. As examples, we micropatterned the protein bovine serum albumin (BSA) and the enzyme horseradish peroxidase (HRP). The catalytic activity of HRP was shown to be retained after being cross‐linked to the support. When coupled with solid‐phase peptide synthesis, the technique allowed synthetic peptides to be constructed in patterns again having micrometer dimensions. Synthetic polypeptides, polylysine, were constructed in patterns with dimensions that approached the practical limit of resolution for optical lithography at 1–2 μman. The patterns of immobilized molecules and synthetic peptides were visualized using histochemical methods together with light and fluorescence microscopy. The protein and peptide patterning technique described here is an advance in the field of bioelectronics. In particular, it should now be possible to devise novel methods for interfacing with biological systems and constructing new devices for incorporation into miniaturized biosensors.
Spin Labeling and Kinetic Studies of a Membrane‐Immobilized Proteolytic Enzyme
Authors: Zhuangt Ping , Butterfield D. Allan
Published: March 2005

Membrane‐based bioreactors can greatly influence the rate and extent of chemical reactions and consequently lower the costs associated with the corresponding engineering processes. However, in order to progress in this area, greater understanding of the relationship of the structure and function of bioreactor systems is required. In this study, a proteolytic enzyme, papain (EC 3.4.22.2), was covalently coupled onto the surface of a vinyl alcohol/vinyl butyral copolymer (PVB) membrane employing either glutaraldehyde (GA) or l,l'‐carbonyldiimidazole (CDI). Various kinetic and performance properties of the immobilized papain were studied. It was found that these characteristics of the membrane‐bound papain depended on the immobilization method. The CDI‐immobilized papain bioreactor was used, although the apparent Michaelis constant, Km, of the CDI‐immobilized papain was larger than that of the GA‐immobilized enzyme. In separate experiments, a six‐carbon spacer was also used between the membrane support and the covalently‐linked enzyme. It was found that the insertion of the spacer reduced the disturbance of the enzyme system, resulting in a decreased Km, which was now closer to the value for the free enzyme. Electron paramagnetic resonance (EPR) techniques of spin labeling were used for the first time to examine the conformational change and the active site structure of an enzyme covalently immobilized to a membrane. The structural changes of the active site of papain upon immobilization with and without a spacer were in agreement with the functional properties of the enzyme.
Activity of Thiolsubtilisin in Organic Solvents
Authors: Chatterjee Sudipta , Russell Alan J.
Published: March 2005

In this paper, we compare the activities of subtilisin and thiolsubtilisin in anhydrous organic solvents and in mixtures of water and miscible organic media. The information obtained indicates that changing from one organic solvent to another does not significantly affect the stability of the transition state for the reaction. Thus, since the transition state is charged, it is likely that the active site of the enzyme is electrostatically protected from bulk solvent. Further, since the activities of subtilisin and thiolsubtilisin are different in the solvents used, it is clear that the active site serine is a catalytically important residue in both water and organic solvents.
An Anionic Galactomannan Polysaccharide Gum from a Newly‐Isolated Lactose‐Utilizing Bacterium. II. Fermentation Kinetics and Lactose Transport
Authors: Flatt James H. , Cooper Timothy A. , Gonzalez Juan M. , Dogger Dina E. , Lightfoot Edwin N. , Cameron Douglas C.
Published: March 2005

The facultative bacterium Erwinia ATCC 55046, isolated from the soil of a wheytreated farm field, produces an anionic galactomannan polysaccharide gum (designated lactan gum) from lactose and other sugars. On lactose semidefined medium in a 5‐L fermentor, the organism had a maximum growth rate of 0.66 h−1 and produced 26 g/L polysaccharide with a yield on lactose of 0.66 g/g and a volumetric productivity of 0.6 g/(L.h). In shake flasks, the best final gum concentration was obtained with a carbon to nitrogen ratio of 40:1 (w/w). Gum yield and concentration was greater on disaccharides than on monosaccharides. The organism was found to transport lactose by a permease system as opposed to a phosphotransferase system. Lactan gum can be produced efficiently from lactose, the major sugar in whey and whey permeate.
A Fiber‐Optic System for Measuring Single Excitation‐Dual Emission Fluorescence Ratios in Real Time
Authors: Mccarthy John F. , Magin Richard L. , Kisaalita William S. , Slininger Patricia J.
Published: March 2005

The development and subsequent evaluation of a fiber‐optic system for measuring single excitation—dual emission fluorescence ratios in real time is described. The design of the flashlamp excitation source, optics, electronics, and computer software is discussed. The dual emission pH sensitive fluorophore 1, 4‐dihydroxyphthalonitrile (1, 4‐DHPN) was used to test the performance of this system. The flexible design of this modular system permits the use of other single excitation—dual emission fluorophores by simply changing the appropriate optical filters. Upon a single 340–380‐nm excitation, pH‐sensitive emissions were monitored at 488 nm and 434 nm. The ratio of these emissions (488/434) was then computed in real time, for a 2 mM solution of 1, 4‐DHPN, while the pH was titrated over the range 5–9. The nonlinear, system‐dependent, calibration curve of pH versus the ratio of emission wavelengths was empirical fit by a fourth‐order polynomial (r2 = 0.995). Reliable pH measurements in the range 6–8 were obtained using concentrations of 1, 4‐DHPN as low as 50 μM. The standard deviation of pH measurements using a 1 mM solution of 1, 4‐DHPN, near neutral pH, was found to be approximately 0.1 pH unit.
Electrokinetic Isolation of Vesicles and Ribosomes Derived from Serratia marcescens
Authors: Hawker Debra T. L. , Todd Paul , Davis Robert H. , Lawson Robert C. , Rudge Scott R.
Published: March 2005

Ribosomes and vesicles derived from the bacterium Serratia marcescens were separated from each other and from solubles using density gradient electrophoresis. Transport relationships were used to determine the electrophoretic mobilities of the particles. The effects of convection, sedimentation and diffusion were found to be negligible. The electrophoretic mobility obtained for the ribosome peak is –7 × 10–5 cm2/(V.s). Under appropriate conditions, two vesicle peaks were obtained, the first with a mobility of –4 × 10–5 cm2/(V.s) and the second with –9 × 10−6 cm2/(V.s). This information can be used to predict the resolution of the separands in large‐scale electrophoretic separations.
Techniques for Studies on Growth Characteristics of Human Prostatic Cancer Cells
Authors: Bauer Johann , Grimm Daniela , Hofstaedter Ferdinand , Wieland Wolf
Published: March 2005

The methods described in this article seem to be useful for studies on the growth characteristics of malignant epithelial prostate cells which are still under the influence of healthy cells. Before establishing primary cultures, pieces of tissue were sectioned in such a way that they could be unfolded to obtain a large surface. These pieces were treated enzymatically and then incubated for at least 4–6 weeks. In this time, cells grew or migrated out of the tissue and spread over the surface of the culture flasks. The viable single cells harvested from these primary cultures were characterized flow cytometrically, fractionated by countercurrent centrifugal elutriation, or further incubated above agarose so that they formed three‐dimensional spheroids. Cytometric determinations of cellular cytokeratin, vimentin, and DNA were performed before and after incubation. They suggested that the percentages of cytokeratin‐positive (epithelial) cells, vimentin‐positive cells (fibroblasts), and aneuploid cells remained at levels (in vitro) similar to those within the pieces of tissue used for the culturing experiments, respectively. Since our culture technique allows the propagation of human epithelial prostate cells in vitro as they would grow in vivo under the control of the surrounding tissue, the method should help to investigate which particular treatment of the cells influences the growth of the malignant cells, while they are still surrounded by other cells of the same prostatic organ.
Cation‐Exchange Displacement Chromatography of Proteins with Protamine Displacers: Effect of Induced Salt Gradients
Authors: Gerstner Joseph A. , Cramer Steven M.
Published: March 2005

Protamine was investigated for its utility as a protein displacer in cation‐exchange systems. Although the protamine solution contained several variants of the molecule, the high affinity of all of the components in this heterogeneous biopolymer enabled it to act as an efficient protein displacer. To facilitate parameter estimation of the protamine, a preliminary purification was carried out by preparative elution chromatography. Chromatographic parameters of both the feed proteins and protamine displacer were obtained for use in a multicomponent steric mass action ion‐exchange displacement model. Model simulations were compared to displacement results under both moderate and intense induced salt gradient conditions. In both cases, excellent agreement was obtained between the displacement experiments and theoretical predictions. In addition, these studies serve to dramatize the importance of induced salt gradients in ion‐exchange displacement systems.
One‐Step Purification of a Model Periplasmic Protein from Inclusion Bodies by Its Fusion to an Effective Metal‐Binding Peptide
Authors: Beitle Robert R. , Ataai Mohammad M.
Published: March 2005

It has been demonstrated that the addition of a metal‐binding amino acid sequence to an exposed terminus of a protein can be useful for purification using immobilized metal affinity chromatography (IMAC). Polyhistidine extensions, wherein sequential histidyl residues are placed at the end of a protein, have been utilized for protein purification through IMAC. Natural metal‐binding peptides may also serve as starting points for the design of an affinity tail. As a model system, an octapeptide derived from angiotensin I was fused to TEM‐β‐lactamase. When the modified protein was expressed in Escherichia coli, a one‐step purification of this recombinant protein was accomplished from resolubilized inclusion body material.
Compressive Characteristics of Freeze‐Dried Agar and Alginate Gel Sponges
Authors: Nussinovitch A. , Velez‐Silvestre R. , Peleg M.
Published: March 2005

Mechanically stable solid sponges can be produced by freeze drying 2 % agar and 1 % alginate gels. The sponges have characteristic compressive stress‐strain curves that can be described by a three‐parameter model originally developed for polymeric sponges and bakery products. Internally produced bubbles, formed by immersing bicarbonate‐containing gels in an acid bath, resulted in a considerable loss of mechanial integrity in the dry agar sponges but not in those of the alginate.
Increased Activity of Chromobacterium viscosum Lipase in Aerosol OT Reverse Micelles in the Presence of Nonionic Surfactants
Authors: Yamada Yasushi , Kuboi Ryoichi , Komasawa Isao
Published: March 2005

Modification of AOT reverse micellar systems with various alkyl glucosides and nonionic surfactants was attempted in order to improve the activity to Chromobacterium viscosum lipase. Tweens and Tritons, having the poly(oxyethylene) chain, have been shown to be solubilized at the oil—water interface of AOT micelles. Alkyl glucosides were also solubilized at the interface. These additives form mixed micelles with AOT and increase the concentration of the micelles. In contrast, Spans were found to be solubilized in the micro water pool of AOT micelles. Addition of nonionic surfactants decreased the hydrophobicity of the reverse micellar systems. Improvement of the lipase activity was achieved by the addition of nonionic surfactants, because of suppression of the hydrophobic and electrostatic interaction between AOT and lipase. The addition of nonionic surfactant having a poly(oxyethylene) chain has been shown to effectively increase the activity of the lipase in reverse micelles.
Effect of Silicone Antifoam on Shear Sensitivity of Hybridoma Cells in Sparged Cultures
Authors: Van Der Pol Leo A. , Bonarius Dick , Van De Wouw Geert , Tramper Johannes
Published: March 2005

Addition of high concentrations of silicone antifoam (AF) to a suspension of hybridoma cells in a bubble column reduces the death rate when using medium without a protective component. In contrast, high AF concentrations can cause detrimental effects in a sparged medium with a protective component, such as is applied for large‐scale animal cell cultures.
Evaluation of Crude Hydroxypropyl Starch As a Bioseparation Aqueous‐Phase‐Forming Polymer
Authors: Venǎncio Armando , Teixeira José A. , Mota Manuel
Published: March 2005

A new aqueous two‐phase system based on crude hydroxypropyl starch (HPS) and poly(ethylene glycol) (PEG) is described. Unlike other polymer‐polymer aqueous two‐phase systems, the HPS used is commercially available. Previous works have characterized almost exclusively purified carbohydrates or their derivatives, having neglected the large amount of gums and carbohydrates already used in the paper, food, or textile industries. Two‐phase aqueous polymer systems based on such commercial polymers may have as a drawback their degree of purity, which may affect the separation procedure. However, their lower cost should encourage studies on their use in aqueous two‐phase polymer extraction. The influence of temperature, pH, and ionic strength on the partition of several different molecular weight and isoelectric point proteins and whole cells was studied. The results obtained were compared with corresponding dextran‐PEG systems, Aquaphase PPT‐PEG systems, crude dextran‐PEG systems, maltodextrin‐PEG systems, and Klucel L‐pluronic systems. Protein partition coefficient results exhibited higher values than those in corresponding systems. It is shown that this system, based on a crude hydroxypropyl starch, is highly attractive as an initial purification step.
Dissolved Carbon Dioxide Measurement and Its Correlation with Operating Parameters in Fermentation Processes
Authors: Dahod Samun K.
Published: March 2005

A silicone tubing probe was used to measure dissolved carbon dioxide in two industrial fermentation processes. The two processes differed markedly in their broth viscosities: one had high viscosity (2000–3000 cP) and the other had low viscosity (<100 cP). The goal was to study changes in dissolved carbon dioxide concentration in fermentation broth when operating parameters, such as total pressure, air flow rate and, agitation rate, were modified. It was shown that in high‐viscosity fermentation broths the dissolved carbon dioxide concentration can reach as high as 190% of that calculated by assuming an equilibrium between the air coming out of the fermentor and the fermentation broth. For low‐viscosity broth, the highest dissolved carbon dioxide concentration was found to be about 133 % of the equilibrium value. On the basis of the material balance over the entire fermentor, the ratio of actual dissolved carbon dioxide partial pressure and carbon dioxide partial pressure in air leaving the fermentor was correlated to the operating parameters of pressure, air flow, and carbon dioxide transfer coefficient. Since carbon dioxide transfer coefficients are not readily available, we recommend a method of using oxygen transfer coefficients to develop a correlation that can be used on‐line to predict changes in dissolved carbon dioxide concentration when operating parameters are changed or when process improvements and scale‐up are undertaken.
Production of Human Alkaline Phosphatase, a Secreted, Glycosylated Protein, from a Baculovirus Expression System and the Attachment‐Dependent Cell Line Trichoplusia ni BTI‐Tn 5B1‐4 Using a Split‐Flow, Air‐Lift Bioreactor
Authors: Chung I. S. , Taticek R. A. , Shuler M. L.
Published: March 2005

A split‐flow, air‐lift bioreactor for the cultivation of insect cells to produce recombinant protein is described. It can be used advantageously with attached cell systems. This bioreactor incorporates two sections: a rise and a downcomer. Trichoplusia ni BTI‐Tn 5B1‐4 cells are grown on a support material of glass beads or microcarriers placed in the downcomer. This cell line is more productive than other commonly used insect cell lines, but it has the disadvantage of being difficult to use at large volumes since it is not easily adaptable to suspension culture. Adequate oxygen demand is supplied by sparging without direct exposure of cells to air bubbles. Nutrients are supplied convectively to the attached cells on support material as the fluid flows through the downcomer. The split‐flow, air‐lift bioreactor appears to be suitable for insect cell culture and is potentially scalable. It can provide a high surface‐to‐volume ratio and can be operated in batch or continuous mode. A lab‐scale prototype bioreactor has been constructed and tested for the production of a secreted, glycosylated recombinant protein (human alkaline phosphatase) or seAP using an Autographa californica multiple nuclear polyhedrosis virus (AcMNPV) vector. With a ratio of riser cross‐sectional area to downcomer cross‐sectional area of 1, an aspect ratio of 4.4, an air‐flow rate of 54 mL/min in the riser, and a bed of 2400 3‐min nonporous glass beads, 10.7 μg/mL of seAP was produced using an MOI (multiplicity of infection or the ratio of plaque‐forming units to cells) of 10. On a specific productivity basis, the split‐flow, air‐lift system and T‐flasks gave similar values (about 13 μg/106 cells).
Ethanol Production in Baker's Yeast: Application of Experimental Perturbation Techniques for Model Development and Resultant Changes in Flux Control Analysis
Authors: Schlosser Paul M. , Riedy Gerard , Bailey James E.
Published: March 2005

Recent theoretical results (Schlosser and Bailey, 1990; Schlosser et al., 1993) suggest that it is possible to model elements of metabolism to the extent necessary for flux control analysis using only the results of perturbation experiments. In particular, the development of detailed, mechanistic descriptions of certain relevant processes (which would otherwise require prohibitive effort) can be avoided by direct use of experimental results. Anaerobic glycolysis in Saccharomyces cerevisiae AMW‐13C was studied both under unperturbed conditions and under several experimental perturbations. Changes in pathway fluxes and metabolite levels relative to the unperturbed values were observed and analyzed to determine the relationships that exist between several of these quantities. These relationships were used to refine and extend a previously determined model for the glycolytic pathway (Galazzo and Bailey, 1990, 1991). The refined model was then used to study the flux control characteristics of the pathway. The analysis indicates that the reaction catalyzed by phosphofructokinase (PFK), while retaining significant flux control, is less important than previous studies suggested and that glucose uptake is the predominant rate‐controlling step when small changes are considered. The model indicates that larger increases in the amount of any single pathway enzyme yield no more than a 36% increase in ethanol production.
Intermolecular Electrostatic Interactions and Their Effect on Flux and Protein Deposition during Protein Filtration
Authors: Palecek Sean P. , Zydney Andrew L.
Published: March 2005

Although membrane filtration is used extensively to process protein solutions containing a variety of electrolytes, there is currently little fundamental understanding of the effect of the solution environment (and in particular, the solution pH) on the filtrate flux in these systems. We have obtained data for the flux and sieving coefficients during the batch (stirred cell) filtration of solutions of bovine serum albumin, immunoglobulins, hemoglobin, ribpnuclease A, and lysozyme through 0.16‐μm micro‐filtration membranes at different pH values. The flux declined significantly for all five proteins due to the formation of a protein deposit on the upper surface of the membrane. The quasi‐steady ultrafiltrate fluxes at the individual protein isoelectric pH's were essentially identical, despite the large differences in molecular weight and physico‐chemical characteristics of these proteins. The flux increased at pH's away from the isoelectric point, with the data well‐correlated with the protein surface charge density. These results were explained in terms of a simple physical model in which the protein deposit continues to grow, and thus the flux continues to decline, until the drag force on the proteins associated with the filtrate flow is no longer able to overcome the intermolecular repulsive interactions between the proteins in the bulk solution and those in the protein deposit on the surface of the membrane.
Use of Binding Measurements To Predict Elicitor Dosage Requirements for Secondary Metabolite Production from Root Cultures
Authors: Singh Gurmeet , Reddy G. Ramakrishna , Curtis Wayne R.
Published: March 2005

The strength of binding of a fungal elicitor derived from Rhizoctonia solani to Agrobacterium‐transformed root cultures of Hyoscyamus muticus is assessed. Two techniques are used to estimate the equilibrium binding constant: (1) a dose response, which involves the measurement of sesquiterpene formation in the presence of different amounts of elicitor, and (2) elicitor‐contacting experiments in which the root tissue is exposed to the elicitor for a short period of time and then the unbound elicitor concentration is determined by a bioassay. The equilibrium binding constants estimated by these two techniques were on the same order of magnitude. The value predicted from the dose response technique was 6.44 (mmol of glucose equiv/L of medium)−1, while that obtained from elicitor contact was 1.89 (mmol of glucose equiv/L of medium)−1. The higher binding constant measured by dose response may result from feedback inhibition of secondary metabolite formation at higher elicitor dosages. The magnitude of the binding'constant indicates that elicitor dosage is dependent upon both the amount of tissue in the reactor and the amount of medium present to dilute the added elicitor. Since proposed reactor configurations for plant root culture vary from 200 to 2000 g of tissue per liter of medium, elicitor dosage requirements will vary significantly depending upon the reactor operational conditions. This influence of elicitor binding on reactor design and operation is discussed.
Acid Hydrolysis of K‐Carrageenan in a Batch Reactor: Stochastic Simulation of Change of Molecular Weight Distribution with Time
Authors: Singh S. K. , Shen B. C. , Chou S. T. , Fan L. T.
Published: March 2005

The degradation of K‐carrageenan by acid hydrolysis in a LiCl/HCl (pH 2) buffer solution has been studied at various temperatures in a batch reactor. The reduction in the weight‐average molecular weight and the variation in the molecular weight distribution (MWD) have been followed over time by an SEC‐MALLS‐RI chromatography system. While a deterministic rate equation can be fit to the average molecular weight‐time data, this is not the case for the MWD‐time data. Since depolymerization by hydrolysis usually proceeds via nonspecific (random) bond cleavage, a stochastic approach can be appropriate for the resultant MWD‐time data. In the present work, the master equation has been derived from the stochastic population balance for this batch hydrolysis system characterized by a set of discrete states, each representing a particular molecular weight range. The governing equations for the means, variances, and covariances of the random variables, each representing the number of carrageenan molecules in an individual state, have been extracted from the master equation. The equations for the means have been applied to the MWD‐reaction time data, and the unknown reaction rate parameters have been estimated. These parameters can be used to generate the MWD at any hydrolysis reaction time from any initial MWD. The applicability, limitations, advantages, and sources of error of the derived model have been discussed.
Study of Nitrate Metabolism of Escherichia coli Using Fluorescence
Authors: Trivedi Hiren K. , Ju Lu‐Kwang
Published: March 2005

Public concern about environmental nitrate contamination has increased significantly. To investigate the nitrate metabolism of Escherichia coli, profiles of NAD(P)H fluorescence responding to nitrate/nitrite additions to anaerobic E. coli cultures at the resting or minimum‐growth state were monitored by an on‐line fluorometer. Two different types of fluorescence profiles were observed. Upon nitrate addition, the fluorescence signal dropped instantaneously, indicating the presence of NAD(P)H‐dependent nitrate/nitrite reductases, without previous nitrate/nitrite induction. The signal remained at that level until all of the nitrate was reduced. In some cases, the signal then directly rose back to the earlier anaerobic level. In others, it partially recovered to an intermediate level, remained at that level for a period much longer than the first stage, and then returned to the anaerobic level. The single‐stage response implied that the nitrite formed by nitrate reduction was totally converted to ammonium simultaneously. The two‐stage response, however, resulted from intermediate nitrite accumulation: the rise in fluorescence at the end of the first stage signified nitrate depletion, and the second stage corresponded to reduction of the nitrite accumulated in the first stage. This is supported by the specific rates of nitrate and nitrite reduction determined from the fluorescence profiles. While the former (0.0138 ± 0.0026 g of NO3−‐N/(g of cells)‐h) was about the same in all the runs conducted, the latter was found to be 1 order of magnitude lower than the former in cultures showing the two‐stage response. The presence of ammonium, at 0–50 ppm NH4+‐N, was found to have no appreciable effect on the nitrate reduction rate for the E. coli studied.
Antigen‐Antibody Binding Kinetics for Biosensors: The Fractal Dimension and the Binding Rate Coefficient
Authors: Sadana Ajit
Published: March 2005

The diffision‐limited binding kinetics of antigen (or antibody or substrate)in solution to antibody (or antigen or enzyme) immobilized on a biosensor surface is analyzed within a fractal framework. The change in the fractal dimension, Df, is in the same direction as that in the forward binding rate coefficient, k1. An increase or decrease in the fractal dimension, Df leads to an increase or decrease in the forward binding rate coefficient, k1 respectively, irrespective of whether the antigen or the antibody is immobilized on the biosensor surface. For example, an increase in the concentration of IgG (MRC OX‐19) (antigen) in solution from 1 to 10 μg/mL bound to polyclonal sheep anti‐mouse IgG (antibody) immobilized on immunomagnetic beads leads to decreases in Df from 1.93 to 0.517 and in k1 from 2.72 to 0.064, respectively. The different examples analyzed and presented together indicate one means by which the forward binding rate coefficient, k1, may be controlled, that is, by changing the fractal dimension or the disorder on the surface. The versatility of the analysis is indicated by applying it to different examples for both types of systems, wherein either the antigen or antibody is immobilized on the surface. The analysis should assist in improving the stability, sensitivity, and response time of biosensors.
Batch and Semicontinuous Aggregation and Sedimentation of Hybridoma Cells by Acoustic Resonance Fields
Authors: Pui Phylis W. S. , Trampler Felix , Sonderhoff Stefan A. , Groeschl Martin , Kilburn Douglas G. , Piret James M.
Published: March 2005

Ultrasound was used to enhance the sedimentation of hybridoma cells from medium in a 75 mL resonator chamber. Forces in the acoustic standing waves aggregated the cells, and the aggregates were then rapidly sedimented by gravity. Cell separation increased with acoustic treatment time and cell concentration. The separation efficiency was over 97% for cell concentrations between 1066 and 107 cells/mL. During acoustic treatment at 180 W/L, the medium temperature increased at a rate of 1.3 °C/min. Ultrasonic exposures up to 220 W/L did not influence the viability or subsequent growth and antibody production of the cells. A decrease in cell viability was observed at a power level of 260 W/L. Batch separation efficiencies were as high as 98%. Acoustic separation was tested under semicontinuous operation, and above 90% separation efficiency was achieved at a flow rate of 0.7 L/h.
Microbial Polysaccharides of Applied Interest: Ongoing Research Activities in Europe
Authors: Crescenzi Vittorio
Published: March 2005

After an introductory note on the main reasons underlying the interest in microbial polysaccharides as subjects of fundamental structural/physicochemical investigations, as well as of applied studies, and after a brief historical account, a survey is offered of ongoing research activities in Europe in the broad field of carbohydrate polymers of microbial origin. Attention is drawn to glycopolymers of actual or potential industrial relevance.
Characterization of a Bench‐Scale System for Studying the Biodegradation of Organic Solid Wastes
Authors: Tseng David Y. , Chalmers Jeffrey J. , Tuovinen Olli H. , Hoitink Harry A. J.
Published: March 2005

A bench‐scale system was developed that through its use contributes toward a more fundamental understanding of the composting process. The computer‐controlled system maintained temperature and moisture levels within narrow ranges in thin layers of compost positioned on nylon mesh trays within the vessel. This system minimized complications caused by gradients observed in earlier systems. Specific O2 consumption and CO2 evolution rates were determined on the basis of dynamic mass balances. Water was introduced intermittently. Microbial biomass (ATP) was monitored at specific time intervals throughout the process. The system allows close observation of decomposition of feed materials at constant temperature and moisture levels throughout the process.
Use of an Acrylic Ester Sorbent for the Selective Adsorption of Avermectins
Authors: Chaubal Mahesh V. , Payne Gregory F.
Published: March 2005

Previous work has shown that simple model solutes differing in their abilities to form hydrogen bonds can be selectively adsorbed from a nonpolar solvent onto an acrylic ester sorbent (Amberlite XAD‐7). The goal of this work was to determine whether these sorbents could be used for the selective adsorption of members of the avermectin family of secondary metabolites. When the individual avermectins were studied, avermectin B2a, which has four hydroxyl groups, was observed to adsorb from hexane with a higher affinity than avermectins A2a or B1a, which have only three hydroxyl groups each. When equimolar mixtures of the three avermectins were studied, it was observed that avermectin B2a was preferentially adsorbed from hexane with separation factors of 2.2 × 0.2 and 1.8 × 0.2 relative to avermectins A2a and B1a, respectively.
Metabolic Engineering of Escherichia coli To Enhance Recombinant Protein Production through Acetate Reduction
Authors: Aristidou Aristos A. , San Ka ‐Yiu , Bennett George N.
Published: March 2005

Genetic and metabolic engineering provide powerful and effective tools for the systematic manipulation and fine tuning of cellular metabolic activities. In this study, successful application of such techniques to enhance recombinant protein production by reducing acetate accumulation in Escherichia coli is presented. The alsS gene from Bacillus subtilis encoding the enzyme acetolactate synthase was introduced into E. coli cells using a multicopy plasmid. This newly introduced heterologous enzyme modifies the glycolytic fluxes by redirecting excess pyruvate away from acetate to acetolactate. Acetolactate is then converted to a nonacidic and less harmful byproduct acetoin, which appears in the broth. Furthermore, comparative fermentation studies show that the reduction in acetate accumulation leads to a significant improvement of recombinant protein production. The expression of a model recombinant CadA/β‐galactosidase fusion protein, under the control of a strong pH‐regulated promoter, was found to increase by about 60% for the specific protein activity (to a level of 30% of total cellular protein) and 50% in terms of the volumetric activity in a batch fermenter. In fed‐batch cultivation, the engineered strain achieved a volumetric recombinant protein yield of 1.6 million units/mL (about 1.1 g/L of β‐galactosidase), which represented a 220% enhancement over the control strain. In the meantime, acetate excretion was maintained below 20 mM compared with 80 mM for the control, and the final cell density was improved by 35%.
Conversion of Mixed Waste Office Paper to Ethanol by Genetically Engineered Klebsiella oxytoca Strain P2
Authors: Brooks T. A. , Ingram L. O.
Published: March 2005

Unsorted, mixed waste office paper (MWOP) is an excellent substrate for conversion into fuel ethanol using a recombinant strain of Klebsiella oxytoca which ferments cellobiose and cellotriose to ethanol at near theoretical yields, eliminating the need for supplemental β‐glucosidase. This organism was tested with commercial fungal cellulase in optimized simultaneous saccharification and fermentation experiments (SSF) using MWOP as a substrate (pH 5–15.2, 35 °C). Similar rates and yields were obtained with dilute acid‐pulped (hydrolysis of hemicellulose) and water‐pulped MWOP on a dry weight basis although viscosity was reduced by the acid pretreatment. In simple batch fermentations, 40 g/L ethanol was produced after 48–72 h with 100 g/L MWOP and 1000 filter paper units (FPU) of cellulase/L, a yield of 550 L of ethanol/metric ton. Cellulase usage was further reduced by recycling SSF residues containing bound enzymes in multistage fermentations. This approach reduced the requirement for fungal cellulase while retaining rapid ethanol production and high ethanol yield. In our optimal design, broths containing an average of 39.6 g/L ethanol were produced in three successive stages with an average fermentation time of 80 h (567 FPU of fungal cellulase/L; 6.1 FPU/g of substrate). This represents a yield of 0.426 g of ethanol/g of substrate, 539 L/metric ton, 129 gal/U.S. ton. MWOP contains approximately 90% carbohydrate. Thus the combined efficiency for saccharification and fermentation to ethanol was 83.3% of the theoretical maximum.
Link Between Vasopressin Receptor AVPR1A Promoter Region Microsatellites and Measures of Social Behavior in Humans
Authors: Bachner-Melman Rachel , Zohar Ada H. , Bacon-Shnoor Naomi , Elizur Yoel , Nemanov Lubov , Gritsenko Inga , Ebstein Richard P.
Published: March 2005

Abstract. Two markers near the vasopressin receptor (AVPR1A) gene located on chromosome 12q14-15 were tested for linkage to two complex social behaviors in humans: Sibling relationships and self-presentation style. Self-report questionnaires were administered to 552 same-sex siblings from 248 families. Suggestive linkage was observed between both microsatellites (RS1 and RS3) and the Sibling Relationship Questionnaire Conflict scale (RS1: χ2 = 13.65, LOD = 2.96, p = .0001; RS3: χ2 = 14.54, LOD = 3.16, p = .00007) and the Concern for Appropriateness Scale Self-presentational style (RS1: χ2 = 8.25, LOD = 1.79 p = .002; RS3: χ2 = 8.81, LOD = 1.91, p = .002. The current results provide the first provisional evidence that the vasopressin receptor mediates social behavior in humans and links a specific genetic element to perceived sibling interactions.
Shortwave aerosol radiative forcing over cloud‐free oceans from Terra: 1. Angular models for aerosols
Authors: Zhang Jianglong , Christopher Sundar A. , Remer Lorraine A. , Kaufman Yoram J.
Published: March 2005

Using multiple satellite instruments, we demonstrate a new empirical method for obtaining shortwave (SW) aerosol angular distribution models (ADMs) over cloud‐free oceans. We use nearly a year's worth of multispectral Moderate Resolution Imaging Spectroradiometer (MODIS) data to obtain aerosol properties within a Clouds and Earth Radiant Energy System (CERES) footprint and Special Sensor Microwave Imager (SSM/I) data to obtain near surface wind speed. The new aerosol ADMs are built as functions of near‐surface ocean wind speed and MODIS aerosol optical depth at 0.55 μm (τ0.55). Among the new features are ADMs as a function of the ratio of fine mode to total aerosol optical depth (η), which contains information on aerosol type, and the combination of the CERES rotation azimuth plane scan mode CERES data and MODIS aerosol products to characterize aerosol properties over glint regions. The instantaneous aerosol forcing efficiencies (SW flux per unit optical depth at τ0.55) are 80.5, 63.1, and 73.0 Wm−2, derived using the Earth Radiation Budget Experiment (ERBE), Tropical Rainfall Measuring Mission (TRMM), and the current Terra ADMs, respectively. This study highlights the necessity for building empirical aerosol ADMs as a function of wind speed, τ0.55 and η, and gives examples of newly constructed aerosol ADMs over cloud‐free oceans. We conclude that an overall uncertainty of 10% will be introduced in the derived SW aerosol direct forcing over cloud‐free oceans if the ADMs are constructed without considering aerosol darkening effect over glint regions and aerosol brightening over nonglint regions (like ERBE ADMs) or the variations in aerosol properties (like TRMM ADMs). In a companion paper (Zhang et al., 2005), these new ADMs are used to calculate the shortwave aerosol radiative forcing over the global oceans.
A mitochondrial pool of sphingomyelin is involved in TNFα-induced Bax translocation to mitochondria
Authors: BIRBES Helene , LUBERTO Chiara , HSU Yi-Te , EL BAWAB Samer , HANNUN Yusuf A. , OBEID Lina M.
Published: March 2005

We recently showed that targeting bSMase (bacterial sphingomyelinase) specifically to mitochondria caused accumulation of ceramide in mitochondria, and induced cytochrome c release and cell death [Birbes, El Bawab, Hannun and Obeid (2001) FASEB J., 15, 2669–2679]. In the present study, we investigated the role of this mitochondrial pool of ceramide in response to a receptor-mediated event, namely TNFα (tumour necrosis factor α), and the involvement of this mitochondrial pool of ceramide in Bax translocation to mitochondria, an event that precedes cytochrome c release. Treatment of MCF7 cells with TNFα caused an increase in ceramide levels in the mitochondrial fraction which accompanied Bax translocation to mitochondria. Targeting bSMase to mitochondria specifically resulted in Bax translocation to mitochondria, suggesting that the mitochondrial ceramide pool is involved in Bax translocation. Moreover, in a reconstituted cell-free system, treatment of isolated mitochondria with bSMase enhanced Bax association with mitochondrial membranes. Collectively, these results suggest that the generation of ceramide in mitochondria in response to TNFα is sufficient to induce Bax translocation to mitochondria and subsequent cytochrome c release and cell death.
Rotational and vibrational dynamics of ethylene in helium nanodroplets
Authors: Lindsay C. M. , Miller R. E.
Published: March 2005

Rotationally resolved infrared spectra are reported for the asymmetric C–H stretching fundamental bands of C2H4 in helium nanodroplets, as well as two weak combination bands. The J=2 rotor levels are strongly shifted from the energies estimated from a rigid rotor calculation and can be accounted for with two centrifugal distortion constants. The excited states of the three bands with B3u symmetry are strongly coupled in the gas phase and exhibit lifetimes >100ps in helium, with the upper member of the polyad exhibiting the shortest lifetime. In contrast, the ν9 band (B2u symmetry) exhibits very broad, homogeneously broadened line profiles (full width at half maximum ∼0.5cm−1) corresponding to an excited state lifetime of ∼10ps. This short lifetime is presumed to be due to an efficient, solvent mediated vibration-to-vibration relaxation process. In addition, the absence of transitions to the 221 and 220 rotor levels in the ν9 band suggests they form rotational resonances with the elementary modes of helium, resulting in very short excited state lifetimes of less than 2ps.
Polarization resolved Hα spectra from the large helical device: Emission location, temperature, and inward flux of neutral hydrogen
Authors: Iwamae Atsushi , Hayakawa Masayuki , Atake Makoto , Fujimoto Takashi , Goto Motoshi , Morita Shigeru
Published: March 2005

Spectral profiles of the Hα line emitted from the large helical device plasma [O. Motojima et al., Phys. Plasmas 6, 1843 (1999)] have been measured with polarization-separation optics and a high-resolution spectrometer. Besides the underlying high-temperature component, which probably arises from charge-exchange recombination, the profiles are interpreted as superpositions of Zeeman profiles for two different magnetic field strengths. The emission locations are thus identified on the magnetic field map; the emissions are localized in the inner and outer regions just outside the ergodic layer, and each field-strength contribution to the overall Zeeman profile represents two radiator temperatures, and inward atom flow velocities in the range of (1–7)×103m∕s.
The reaction of anthracene with OH radicals: An experimental study of the kinetics between 58 and 470K
Authors: Goulay F. , Rebrion-Rowe C. , Le Garrec J. L. , Le Picard S. D. , Canosa A. , Rowe B. R.
Published: March 2005

The first direct measurement of the reaction rate constant of a polycyclic aromatic hydrocarbon in the gas phase in the temperature range 58–470K is reported. The reaction is OH+ anthracene and the experiment has been performed in a continuous flow Cinétique de Réaction en Ecoulement Supersonique Uniforme apparatus, which had to be modified for this purpose. Pulsed laser photolysis of H2O2 has been used to generate OH radicals and laser-induced fluorescence to observe the kinetic decay of the radicals and hence determine the rate coefficients. The reaction is found to be fast, and the rate constant increases monotonically as the temperature is lowered. The rate coefficients match the expression k(cm3molecules−1s−1)=1.12×10−10(T∕300)−0.46.
Ellipsometric study of adsorption on nanopatterned block copolymer substrates
Authors: Mykhaylyk T. A. , Evans S. D. , Hamley I. W. , Henderson J. R.
Published: March 2005

We report ellipsometrically obtained adsorption isotherms for a carefully chosen test liquid on block copolymer films of Kraton G1650, compared with adsorption isotherms on homogeneous films of the constituent polymers. Standard atomic force microscopy images imply the outer surface of Kraton G1650 is chemically patterned on the nanoscale, but this could instead be a reflection of structure buried beneath a 10 nm layer of the lower energy component. Our test liquid was chosen on the basis that it did not dissolve in either component and in addition that it was nonwetting on the lower energy polymer while forming thick adsorbed films on pure substrates of the higher energy component. Our ellipsometry data for Kraton G1650 rule out the presence of segregation by the lower energy constituent to the outer surface, implying a mixed surface consistent with Cassie’s law. We discuss implications of our findings and related work for the outer surface structures of block copolymer films.
Porous silicon mirrors with enlarged omnidirectional band gap
Authors: Xifré-Pérez E. , Marsal L. F. , Pallarès J. , Ferré-Borrull J.
Published: March 2005

Multilayers consisting of the periodic repetition of two layers with different refractive indices (nH and nL) may have an omnidirectional band gap, the width of which depends on the incidence medium and on the refractive index ratio nH∕nL. In porous silicon, this ratio is limited by the material and fabrication characteristics. This article makes a theoretical study of two multilayer structures in order to enlarge the omnidirectional band gap with the same refractive index ratio by varying only the thickness of the period. These structures are named balanced and unbalanced mirrors and consist of a few periodic multilayers stacked together. Porous silicon balanced and unbalanced mirrors with enlarged omnidirectional band gap for 1.55-μm applications are proposed. The results obtained with these mirror structures are compared with the results obtained with a chirped structure with the same characteristics.
Continuous-wave laser oscillation on the 1315nm transition of atomic iodine pumped by O2(a1Δ) produced in an electric discharge
Authors: Carroll David L. , Verdeyen Joseph T. , King Darren M. , Zimmerman Joseph W. , Laystrom Julia K. , Woodard Brian S. , Benavides Gabriel F. , Kittell Kirk , Stafford D. Shane , Kushner Mark J. , Solomon Wayne C.
Published: March 2005

Laser action at 1315nm on the I(P1∕22)→I(P3∕22) transition of atomic iodine is conventionally obtained by a near-resonant energy transfer from O2(a1Δ) which is produced using wet-solution chemistry. The difficulties in chemically producing O2(a1Δ) has motivated investigations into purely gas phase methods to produce O2(a1Δ) using low-pressure electric discharges. In this letter, we report on the demonstration of a continuous-wave laser on the 1315nm transition of atomic iodine where the O2(a1Δ) used to pump the iodine was produced by a radio-frequency-excited electric discharge. The electric discharge was sustained in a He∕O2 gas mixture upstream of a supersonic cavity which is employed to lower the temperature of the continuous gas flow and shift the equilibrium of atomic iodine in favor of the I(P1∕22) state. The laser output power was 220mW in a stable cavity composed of two 99.99% reflective mirrors.
Direct patterning of organic light-emitting devices by organic-vapor jet printing
Authors: Sun Yiru , Shtein Max , Forrest Stephen R.
Published: March 2005

We demonstrate small molecular weight, fluorescent organic light-emitting devices directly patterned by organic-vapor jet printing (OVJP). In OVJP, a hot inert carrier gas picks up a molecular organic vapor that expands when passed through a nozzle, resulting in physisorption of the molecules onto a cooled substrate. A (1cm)2 flat organic semiconductor thin film is achieved by sweeping the nozzle over the substrate to print closely spaced lines. Using an array of nozzles, multiple parallel stripes are simultaneously printed at a growth rate of 3.4Åcm2∕s. Organic heterostructure fluorescent-light-emitting devices consisting of a broad area hole transport layer, followed by a striped electron transport layer, both printed by OVJP, yield an external quantum efficiency of (0.84±0.03)%, which is comparable to that of similar devices deposited by vacuum thermal evaporation.