bluuebunny/crossref_metadata_2025 huggingface dataset:
Estimation of cell volume and biomass of penicillium chrysogenum using image analysis
Authors: Packer H. L. , Keshavarz‐Moore E. , Lilly M. D. , Thomas C. R.
Published: December 2004

A methodology for the estimation of biomass for the penicillin fermentation using image analysis is presented. Two regions of hyphae are defined to describe the growth of mycelia during fermentation: (1) the cytoplasmic region, and (2) the degenerated region including large vacuoles. The volume occupied by each of these regions in a fixed volume of sample is estimated from area measurements using image analysis. Areas are converted to volumes by treating the hyphae as solid cylinders with the hyphal diameter as the cylinder diameter. The volumes of the cytoplasmic and degenerated regions are converted into dry weight estimations using hyphal density values available from the literature. The image analysis technique is able to estimate biomass even in the presence of nondissolved solids of a concentration of up to 30 gL−1. It is shown to estimate successfully concentrations of mycelia from 0.03 to 38 gL−1. Although the technique has been developed for the penicillin fermentation, it should be applicable to other (nonpellected) fungal fermentations.
Effects of ammonia and lactate on hybridoma growth, metabolism, and antibody production
Authors: Ozturk Sadettin S. , Riley Mark R. , Palsson Bernhard O.
Published: December 2004

The influence of ammonia and lactate on cell growth, metabolic, and antibody production rates was investigated for murine hybridoma cell line 163.4G5.3 during batch culture. The specific growth rate was reduced by one‐half in the presence of an initial ammonia concentration of 4 mM. Increasing ammonia levels accelerated glucose and glutamine consumption, decreased ammonia yield from glutamine, and increased alanine yield from glutamine. Although the amount of antibody produced decreased with increasing ammonia concentration, the specific antibody productivity remained relatively constant around a value of 0.22 pg/cell‐h. The specific growth rate was reduced by one‐half at an initial lactate concentration of 55 mM. Although specific glucose and glutamine uptake rates were increased at high lacatate concentration, they showed a decrease after making corrections for medium osmolarity. The yield coefficient of lactate from glucose decreased at high lactate concentrations. A similar decrease was observed for the ammonia yield coefficient from glutamine. At elevated lactate concentrations, specific antibody productivities increased, possibly due to the increase in medium osmolarity. The specific oxygen uptake rate was insensitive to ammonia and lactate concentrations. Addition of ammonia and lactate increased the calculated metabolic energy production of the cells. At high ammonia and lactate, the contribution of glycolysis to total energy production increased. Decreasing external pH and increasing ammonia concentrations caused cytoplasmic acidification. Effect of lactate on intracellular pH was insignificant, whereas increasing osmolarity caused cytoplasmic alkalinization.
Selective recycle of viable animal cells by coupling of airlift reactor and cell settler
Authors: Hülscher Manfred , Scheibler Uwe , Onken Ulfert
Published: December 2004

A new system for the perfusion culture of animal cells in suspension is described. It consists of an airlift loop reactor and a settling tank for cell retention. Insufficient nutrient and oxygen supply of the cells in the settling tank was prevented by cooling the cell suspension before entering the settler. As a result, the catabolic activity of the cells in the settler was reversibly reduced. Furthermore, the density gradient induced by cooling caused a liquid motion through the settler. Thus, it was not necessary to pump medium containing shear, sensitive cells. With this simple system, it was possible to prduce 2 to 5 g of antibodies in a 5.4‐L reactor in continuous runs of 400 to 600 h. The productivity was increased by a factor of 17 and the cell density was 4 times higher in comparison with the corresponding batch system. The cell retention system was found to have the property of separating viable and nonviable cells. With the increasing perfusion rate, dead cells and debris were preferably washed out. For perfusion rates up to 1.3 d−1, the retention efficiency of the settler was nearly 100% for viable cells; hence, this system may show advantages at the industrial scale.
Protamine immobilization and heparin adsorption on the protamine‐bound cellulose fiber membrane
Authors: Kim Jae‐Seung , Yang Arthur J. , Yang Victor C.
Published: December 2004

Immobilization of protamine to the inner lumen of cellulose hollow fibers has been shown useful in preventing both heparin‐ and protamine‐induced complications during an extracorporeal blood circulation procedure. The current study examined the effects of variables on the immobilization of protamine to cyanogen bromide (CNBr)‐activated cellulose hollow fibers. The degree of protamine immobilization was controlled by three independent parameters: the amount of CNBr used during the activation process, the duration of the coupling process, and the protamine concentration in the coupling solution. By the adjustment of these parameters, cellulose fibers containing desired amounts of immobilized protamine (ranging from 1 to 20 mg of immobilized protamine per gram of dry fibers) were readily prepared.

Heparin adsorption to the protamine‐bound cellulose fibers was also examined. The adsorption isotherm followed a Langmuir adsorption model. The amount of heparin adsorbed was dependent on both the heparin concentration in the substrate solution and the protamine loading on the fibers. The Langmuir adsorption constant K was estimated to be 0.37 ± 0.06 mL/mg, whereas the saturation capacity Qs of the protamine‐bound fibers increased with increasing the protamine loading.
Effects of lactate concentration on hybridoma culture in lactate‐controlled fed‐batch operation
Authors: Omasa Takeshi , Higashiyama Ken‐Ichi , Shioya Suteaki , Suga Ken‐ichi
Published: December 2004

To investigate the effects of lactate on cell growth and antibody production, a new method of maintaining the lactate concentration constant in a fed‐batch culture is described. When the pH was initially adjusted by sodium hydroxide, the specific growth rate decreased and specific death rate increased with an increase of lactate concentration. To investigate whether the inhibition was due to the lactate concentration itself or to the osmotic pressure, the effect of the osmotic pressure adjusted by sodium chloride was compared with that of sodium lactate. When the osmotic pressure was adjusted to same condition as that of sodium lactate using sodium chloride, the specific growth data showed the same degree of growth inhibition. It was thus evident that the inhibition to cell growth was mainly due to osmotic pressure while lactate production from glucose was found to be inhibited by the lactate itself compared with sodium chloride. The specific antibody production rate had a maximum value within a certain range of lactate concentration. Moreover, specific antibody production rate had a unified relationship with the kinetic parameter μ, in spite of the different causes of inhibition by lithium lactate and sodium lactate. A certain “trade‐off” relationship between growth and antibody production existed at higher growth rates.
Mixing in liquid‐impelled loop reactors
Authors: Van Sonsbeek H. M. , Van Der Tuin S. P. , Tramper J.
Published: December 2004

The liquid‐impelled loop reactor is a new column‐type bioreactor. The design of this device is based on the principle of the air‐lift loop reactor. In the external‐loop configuration used in this work, descending perfluorochemical drops bring about circulation of the continuous aqueous phase. Mixing of this continuous phase is characterized per section of the rector. Axial‐dispersion coefficients for the tube with two‐phase flow are determined and correlated with the energy dissipation in the tube. Comparisons with similar systems such as bubble columns and air‐lift loop reactors are made. Overall mixing parameters are derived and used for calculation of the number of circulatins needed to achieve a certain degree of mixing. The hydrodynamic model from previous work is tested for the reactor configurations of this work. It can be useful to calculate circulation times.
Rapid protein release from Escherichia coli by chemical permeabilization under fermentation conditions
Authors: Naglak Thomas J. , Wang Henry Y.
Published: December 2004

Overall protein release greater than 75% in less than 1 h can be attained by exposing exponentially growing Escherichia coli cells to 0.4 M guanidine plus 0.5% Triton X‐100 at 37°C in medium. Cell growth stops immediately upon addition of the chemicals, but the cells are not lysed. Guanidine concentrations lower than 0.2 M, in conjunction with 0.5% Triton X‐100, do not release significant intracellular protein, nor do they inhibit cell growth. Under these conditions, the cells undergo an adaptation that confers resistance to protein release by further treatment with guanidine and Triton X‐100. Cells treated with 0.2 M guanidine plus 0.5% Triton X‐100 display intermediate behavior. Protein release is approximately 35%, and growth is temporarily interrupted by an extended lag phase. Subsequent resumption of cell growth results in resistant cells and no additional protein release. This resistance is shown to be reversible and is most likely due to physiological adaptation rather than genetic mutation.
Production of superoxide dismutase in Streptococcus lactis by a combination of use of hyperbaric oxygen and fermentation with cross‐flow filtration
Authors: Taniguchi Masayuki , Hoshino Kazuhiro , Itoh Takehiro , Kumakura Hiroshi , Fujii Michihiro
Published: December 2004

The production conditions of superoxide dismutase (SOD) in the cells of Streptococcus lactis by using hyperbaric oxygen (O2) are described. The SOD activity of anaerobically grown cells was 5–6 U/mg protein. When the culture broth was pressurized by O2 at 6 atm, the SOD activity was more than twice as high as that under anaerobic culture conditions. However, there is little or no significant increase in SOD activity by exogenous addition of catalase for detoxifying hydrogen peroxide accumulated in the broth and/or controlling the pH of the broth at 6.8 during the pressurization by O2. The increase in SOD activity by hyperbaric O2 was possible not only at the late‐logarithmic growth phase but also at the initial time for the stationary growth phase. For improvement of SOD productivity, we tried a two‐stage culture in which SOD activity in S. lactis cells was enhanced by pressurizing the culture broth using hyperbaric O2 after achievement of a high‐concentration cultivation in the anerobic fermentation system with a microfiltration module.
Effects of temperature and phosphorous concentration on microbial sulfate reduction by Desulfovibrio desulfuricans
Authors: Okabe S. , Characklis W. G.
Published: December 2004

The effects of temperature and phosphorous concentration on the rate and the extent of microbial sulfate reduction with lactate as carbon and energy source were investigated for Desulfovibrio desulfuricans. The continuous culture experiments (chemostat) were conducted at pH 7.0 from 12 to 48°C. The maximum specific growth rate (μmax) was relatively constant in the range 25°C–43°C and dramatically decreased outside this temperature range. The half‐saturation coefficient was minimum at 25°C. Cell yield was highest in the optimum temperature range (35°C–43°C) for growth. Maintenance energy requirements for D. desulfuricans were not significant. Two moles of lactate is consumed for every mole of sulfate reduced, and this stoichiometric ratio is not temperature dependent. Steady state rate and stoichiometric coefficients accurately predicted transient behavior during temperature shifts. The extent of extracellular polymeric substance (EPS) is related to the concentration of phosphorous in the medium. EPS production rate increased with decreased phosphorous loading rate. Failure to discriminate between cell and EPS formation by D. desulfuricans leads to significant overestimates of the cell yield. The limiting C:P ratio for D. desulfuricans was in the range of 400:1 to 800:1.
Liquid‐Phase mass transfer coefficients in bioreactors
Authors: Kawase Yoshinori , Halard Benoit , Moo‐Young Murray
Published: December 2004

Liquid‐phase mass transfer coefficient in bioreactors have been examined. A theoretical model based on the surface renewal concept has been devloped. The predicted liquid‐phase mass transfer coefficients are compared with the experimental data for a mycelial fermentation broth (Chaetomium cellulolyticum) and model media (carboxymethyl cellulose) in a bench‐scale bubble column reactor. The liquid‐phase mass transfer coefficient is evaluated by dividing the volumetric mass transfer coefficient obtained experimentally by the specific surface area estimated using the available correlations. The available literature data in bubble column and stirred tank bioreactors is also used to test the validity of the proposed model. A reasonable agreement between the model and the experimental data is found.
Physico‐chemical features of solvent media in the phases of aqueous polymer two‐phase systems
Authors: Zaslavsky Boris Y. , Borvskaya Anna A. , Gulaeva Nelli D. , Miheeva Larisa M.
Published: December 2004

Solvent polarity and pH in the coexisting aqueous phases of aqueous dextran–poly(ethylene glycol) and dextran–Ficoll two‐phase systems of varied polymer concentrations were examined using the solvatochromic technique and potentiometric measurements, respectively. The relative solvent polarity of the phases, as measured by the solvatochromic technique, is suggested as a measure of the hydration power of water in the phases of aqueous polymer systems. Partitioning of a series of sulphonephthalein dyes in aqueous dextran–poly(ethylene glycol) and dextran–Ficoll two‐phase systems of fixed polymer composition containing 0.01 mol/L universal buffer, pH 7.15, was studied. The results obtained are discussed together with those reported earlier on the physico‐chemical features of aqueous media in the coexisting phases of the systems. It is suggested that the two phases of aqueous polymer systems should be viewed as two immiscible water‐like solvents. The implications of the suggestion for the theoretical treatment of aqueous polymer two‐phase systems are discussed.
Enhancement and regulation of extracellular protein production by Bacillus brevis 47 through manipulation of cell culture conditions
Authors: Wright Charlene P. , Whitney Gordon K. , Daugulis Andrew J. , White Bradley N.
Published: December 2004

Bacillus brevis 47 was cultivated in 2 liter fermentors in semidefined media containing polypeptone with or without glucose or fructose. Neither sugar was essential for growth or extracellular (S‐layer) protein production, and 2.5 to 3.0 g/L protein was accumulated in the medium. When present, glucose was used very slowly, however, fructose was used much more quickly. Dramatic changes in metabolic indicators (dissolved oxygen and pH) were seen when fructose became depleted, and protease was produced, decreasing the amount of protein ultimatelv accumulated in the medium. Using the change in dissolved oxygen as a marker for the time of addition, polypeptone, fructose, or both were used to stimulate protein production. With the addition of polypeptone, on stimulation was achieved, but protease production was suppressed. Addition of fructose did result in a small stimulation of protein production (to 5 g/L) if added once. Further additions resulted in more growth, but no increase in protein production. Various combinations of polypeptone and fructose were also used, with the most effective combination (fructose added early, fructose and polypeptone added later) resulting in an accumulation of 15 g/L protein in the medium. This is comparable to that seen when B. brevis 47 is grown in a complex glucose medium and stimulated with polypeptone addition at 21 hours. These results are discussed with respect to the structure and function of S‐layer proteins, as well as the use of this organism for the production of heterologous proteins.
Storage of solar energy by production of hydrogen peroxide by the blue‐green alga Anacystis nidulans R2: Stimulation by azide
Authors: Morales Inmaculada , Batuecas Santiago , de la Rosa Francisco F.
Published: December 2004

The production of hydrogen peroxide by Anacystis nidulans R2 in presence of methyl viologen occurs by using the redox power from water promoted by the photosystems of the blue‐green alga. Thus, in the presence of the photosynthetic inhibitor DCMU or in the dark, H2O2 production does not take place. In cells permeabilized with lysozyme, the addition of ionophores, which is expected to increase the electron flow, produces only a small increase to initial velocity of hydrogen peroxide production. On the other hand, in nonpermeabilized cells, the addition of superoxide dismutase increases the initial velocity of hydrogen peroxide production, but the net amount accumulated by the system is very low because of posterior decomposition. Preincubation of cells with azide, which inhibits the catalase, prevents the decomposition, thereby increasing drastically the amount of hydrogen peroxide accumulated by the system after a few hours. Hence, H2O2 production appears to be limited mainly because of decomposition by catalase activity rather than by the photosynthetic electron flow rate or the diffusion of products through the cell wall. The net production of hydrogen peroxide by the system was enhanced severalfold by treatment with azide. If one takes into account the use of hydrogen peroxide as fuel due to the large amount of energy released in its dismutation, the photosystem can be a useful tool in the storage of solar energy.
Continuous propionic acid fermentation by immobilized Propionibacterium acidipropionici in a novel packed‐bed bioreactor
Authors: Lewis Vivian P. , Yang Shang‐Tian
Published: December 2004

Continuous propionic acid fermentations of lactate by Propionibacterium acidipropionici were studied in spiral wound fibrous bed bioreactors. Cells were imobilized by natural attachment to fiber surfaces and entrapment in the void volume within the fibrous matrix. A high cell density of ∼37 g/L was attained in the reactor and the reactor productivity was ∼4 times higher than that from a conventional batch fermentation. The bioreactor was able to operate continuously for 4 months without encountering any clogging, degeneration, or contamination problems. Also, the reactor could accept low‐nutrient and low‐pH feed without sacrificing much in reactor productivity. This new type of immobilized cell bioreactor is scalable and thus is suitable for industrial production of propionate. © 1992 John Wiley & Sons, Inc.
A kinetic approach to the thermal inactivation of an imobilized triosephosphate isomerase
Authors: Ábrahám Magdolna , Pénzes Zsolt , Szajáani Béla
Published: December 2004

The process of thermal inactivation of triosephosphate isomerase covalently attached to a silica‐based support activated with p‐benzoquinone was found to be a complex one. At 50°C, a characteristic activation preceding the thermal inactivation was observed. Following the intramolecular changes caused by heat, the values of KM and Vmax were determined during the activation. It was presumed that the complex thermal inactivation kinetics reflects the microheterogeneity of the immobilized enzyme molecules. The phosphate ion proved to be a better stabilizer than the substrate. © 1992 John Wiley & Sons, Inc.
Heparin removal from blood using poly(L‐lysine) immobilized hollow fiber
Authors: Ma Xinghang , Mohammad Syed Fazal , Kim Sung Wan
Published: December 2004

Based on the negative charge density characteristics of heparin, an affinity adsorption technique has been developed for the removal of heparin from blood. Poly(L‐lysine) · HBr (PLL · HBr), a polycation, was immobilized with the help of cyanogen bromide (BrCN) onto poly(ethylene‐vinyl alcohol) (PEVAL) copolymer coated polyethylene (PE) hollow fibers. Heparin bound rapidly onto PLL · HBr imobilized surface in buffer, plasma, and blood. The heparin binding capacity of PLL immobilized surface increased sevenfold as compared to a non‐PLL‐treated control. When heparinized blood was recirculated through a PLL immobilized PEVAL hollow fiber cartridge, the anticoagulant activity of heparin decreased by 85% from initial activity in 25 min. Moreover, circulation of blood through PLL immobilized hollow fiber did not show any adverse effects; no hemolysis was observed and no significant loss of plasma proteins was noted during the heparin removal process. These results suggest that PLL immobilized surface may be utilized for rapid and effective removal of heparin from blood. © 1992 John Wiley & Sons, Inc.
Optimization of accessible catalase activity in polyacrylamide gel‐immobilized Saccharomyces cerevisiae
Authors: Seip John E. , Di Cosimo Robert
Published: December 2004

The permeabilization of Saccharomyces cerevisiae (baker's yeast), either before or after immobilization in polyacrylamide gel (PAG), has been examined as a means to increase the catalase activity of PAG‐immobilized yeast cells. Prior permeabilization of the cells resulted in large losses of catalase activity during immobilization, but permeabilization after immobilization produced increases in the catalase activity of yeast/PAG particles. A dependence of the accessible catalase activity on the concentration of polyacrylamide in permeabilized yeast/PAG particles, and on the method of permeabilization of the immobilized cells, was observed. Optimal levels of stable catalase activity (1000–2000 IU/g PAG particles; ca. 5%–10% of total available activity) were obtained by immobilizing yeast cells (0.5 g wet cells/mL gel) in 10% (w/v) PAG, followed by permeabilization of the entrapped cells with either cetyltrimethylammonium bromide, Triton X‐100 and one freeze‐thaw, or five freeze‐thaw cycles. © 1992 John Wiley & Sons, Inc.
Hydrocarbon degradation by Acinetobacter calcoaceticus RAG‐1 using the self‐cycling fermentation technique
Authors: Brown Wayne A. , Cooper David G.
Published: December 2004

The use of self‐cycling fermentations (SCFs) as a method for dealing with insoluble carbon substrates was examined. The emulsan‐producing Acinetobacter calcoaceticus RAG‐1 was used as the test organism. Limiting concentrations of hexadecane, 1‐hexadecene, or 1‐chlorohexadecane were used as the carbon substrate. The parameters monitored were residual hydrocarbon concentration, cycle time (doubling time), biomass concentration and emulsan concentration. Cycle‐to‐cycle variations of the measured parameters were found to be samll. In all cases, no residual hydrocarbon was detected. The minimum dissolved oxygen concentration was found to correspond with the complete dissappearance of the carbon source. A correlation between minimum dissolved exygen concentration, biomass concentration, and emulsan concentration was noted, thus making it easy to determine when steady‐state conditions had been reached with respect to biomass and emulsan concentrations. The specific emulsan and biomass yields were found to increase during early stages of the fermentation, attaining their respective maxima at steady‐state. Foaming problems often associated with the complete utilization of the insoluble substrate were eliminated using SCF technology, because harvesting occurs immediately following carbon depletion. From the results, SCFs provide a convenient method by which to produce and harvest emulsan. © 1992 John Wiley & Sons, Inc.
Circular dichroism studies on conformational changes in protein molecules upon adsorption on ultrafine polystyrene particles
Authors: Kondo Akihiko , Murakami Fumiyasu , Higashitani Ko
Published: December 2004

The conformational changes in well‐characterized model proteins [bovine ribonuclease A (RNase A), horseradish peroxidase, sperm‐whole myoglobin, human hemoglobin, and bovine serum albumin (BSA)] upon adsorption on ultrafine polystyrene (PS) particles have been studied using circular dichroism (CD) spectroscopy. These proteins were chosen with special attention to molecular flexibility. The ultrafine PS particles were negatively charged and have average diameters of 20 or 30 nm. Utilization of these ultrafine PS particles makes it possible to apply the CD technique to determine the secondary structure of proteins adsorbed on the PS surface. Effects of protein properties and adsorption conditions on the extent of the changes in the secondary structure of protein molecules upon adsorption on ultrafine PS particles were studied. The CD spectrum changes upon adsorption were significant in the “soft” protein molecules (myoglobin, hemoglobin, and BSA), while they were insingnificant in the “rigid” proteins (RNase A and peroxidase). The soft proteins sustained a marked decrease in α‐helix content upon adsorption. Moreover, the native α‐helix content, which is given as the percentage of the α‐helix content in the free proteins, of adsorbed BSA was found to decrease with decreasing pH and increase with increasing adsorbed amount. These observations confirm some well‐known hypotheses for the confirmational chages in protein molecules upon adsorption. © 1992 John Wiley & Sons, Inc.
Performances of aseptic sampling devices for on‐line monitoring of fermentation processes
Authors: Picque D. , corrieu G.
Published: December 2004

Two liquid sampling systems, which are set in the bioreactor and can be sterilized in place, are described. They are composed primarily of an inorganic membrane filter which is either stationary or is set in a rotating motion. These systems have been tested during three types of fermentation processes (ethanol, lactic acid, and polysaccharide productions). The rotating system gave better filtration performances than the stationary sampler which can be used only in alcoholic fermentation. The rotating sampler was coupled with a high‐pressure liquid chromatograph for the on‐line quantitation of the major compounds present in the filtrate. The results are in good agreement with those obtained from off‐line analysis on samples taken manually. © 1992 John Wiley & Sons, Inc.
Application of polystyrene‐bound invertase to continuous sucrose hydrolysis on pilot scale
Authors: Mansfeld J. , Schellenberger A. , Römbach J.
Published: December 2004

Invertase from baker's yeast (Saccharomyces cerevisiae) covalently bound to a macroporous polystyrene anion‐exchange resin via glutaraldehyde was applied to continuous sucrose hydrolysis in packed bed‐reactors. The process was scaled up from 3‐mL laboratory reactors via 0.3‐L reactors to pilot‐scale 50‐L reactors without significant loss of efficiency. The described process allows the production of a wide spectrum of invert sugar syrups with high purity in continuous procedure. The 50‐L reactor was used under process conditions 1 year without significant loss of productivity at a temperature of 40°C. A productivity of 760 g/h was obtained with 1 L invertase–polystyrene complex using a 2.5M sucrose solution as substrate. © 1992 John Wiley & Sons, Inc.
Preparation and application of poly(vinylamine)/alginate microcapsules to culturing of a mouse erythroleukemia cell line
Authors: Wang F. F. , Wu C. R. , Wang Y. J.
Published: December 2004

Poly(vinylamine) was synthesized and used to replace poly‐L‐lysine in forming microcapsule with alginate. Test results indicated that capsules with good mechanical strength and permeability could be obtained under the controlled treatment conditions of poly(vinylamine) and alginate. Application of the current microcapsular system to cell culture was demonstrated by the usage of erythropoietin‐ (EPO‐) producing IW32 mouse erythroleukemia cells. The encapsulated IW32 cells grew to a density of 8 × 107 cells/mL, two times that found in the corresponding poly‐L‐lysine/alginate capsules. The EPO accumulation inside the microcapsule with the current encapsulation system was also higher. A concentration of 7.3 U/mL was attained as compared to 4.3 U/mL in the poly‐L‐lysine/alginate microcapsule. © 1992 John Wiley & Sons, Inc.
Productivity of outdoor algal cultures in enclosed tubular photobioreactor
Authors: Lee Yuan‐Kun , Low Chin‐Seng
Published: December 2004

At quasi‐steady‐state outdoor cyclic fed batch cultures of Chlorella pyrenoidosa, the growth irradiance incidented on the tubular photobioreactor increased about fivefold between 9:00 a.m. and noon. Overheating of the cultures was observed, resulting in decreasing biomass output rate when culture temperature went above 40°C. In cultures with temperature control, the quasi‐steady‐state output rate of all cultures increased throughout the day and leveled off in the late afternoon in high‐density cultures. The daily area output rate was proportional to the density of the cultures. The specific growth rate of the light‐limited cultures increased only marginally (20%) in the morning. © 1992 John Wiley & Sons, Inc.
Methanotrophic attached‐film reactor development and biofilm characteristics
Authors: Fennell Donna E. , Underhill Shelia E. , Jewell William J.
Published: December 2004

The feasibility of using methanotrophs in an attached‐film, fluidized‐bed (MAFFB) reactor system has been under investigation since 1987. Mixed culture, methane‐utilizing attached biofilms were developed on diatomaceous earth particles and on granular activated carbon. The required feed gases, methane and oxygen, were supplied to the attached biofilm in disolved form using separate gas–liquid aeration columns. Biofilm growth was steady despite low influent dissolved methane concentrations (1 to 3 mg/L). A breeder MAFFB operated consistently for 4.1 years with attached biofilm concentrations as high as 51.7 g VS/L static‐bed with minimal biomass wasting and with minimal buffer and nutrient inputs. The maximum biomass concentration observed was 75.6 g VS/L static‐bed in a MAFFB reactor treating trichloroethene. Biofilm thickness reached 160 μm with typical values of 70 μm under methane and oxygen growht‐rate–limited conditions. Biofilm densities of 120 to 190 g VS/L film were observed. Growth rates varied from <0.01/d to 0.17/d. Greater than 90% of the biomass concentration in the bed was attached, and effluent total suspended solids ranged from 5 to 74 mg/L, with an average of 24 mg/L over 27 runs in four MAFFB systems at upflow velocities of 11.4 to 25 m/h. Heterotrophic attached‐film methanotrophs appear to be stable and useful for applications in toxics treatment, and other product manipulations. © 1992 John Wiley & Sons, Inc.
Physical modeling of animal cell damage by hydrodynamic forces in suspension cultures
Authors: Lu George Z. , Gray Murray R. , Thompson B. G.
Published: December 2004

Physical damage of animal cells in suspension culture, due to stirring and sparging, is coupled with complex metabolic responses. Nylon microcapsules, therefore, were used as a physical model to study the mechanisms of damage in a stirred bioreactor and in a bubble column. Microcapsule breaskage folowed first‐order kinetices in all experiments Entrainment of bubbles into the liquid phase in the stirred bioreactor gave more microcapsule breakage. In the bubble column, the bubble bursting zone at gas–liquid interface was primarilu responsible for microcapsule breakage. The forces on the microcapsules were equivalent to an external pressure of approximately 4 × 104 N · m−2, based on the critical microcapsule diameter for survival of 190 μm. A stable foam layer, however, was found to be effective in protecting microcapsules from damage. The microcapsule transport to the gas–liquid interface and entrainment into the foam phase was consistent with flotation by air bubbles. This result implies that additives and operation of bioreactors should be selected to minimize flotation of cells. © 1992 John Wiley & Sons, Inc.
Improved activity retention of enzymes deposited on solid supports
Authors: Wehtje Ernst , Adlercreutz Patrick , Mattiasson Bo
Published: December 2004

Enzymes deposited on solid support usually show good stability when operated in organic solvents. Decreased stability of the enzyme preparations was noticed when low enzyme loadings were used (e.g., with Celite as support; less than 1 mg enzyme/g). It was possible to avoid the activity loss by the addition of an additive which protects the enzyme during the immobilization. Proteins (such as albumin, gelatin, and casein) and poly(ethylene glycol) were effective additives whereas amino acids, monomeric carbohydrates, and polysaccharides had no effect. The amount of additive needed for stabilization was shown to depend on the structure of the support, more additive being required for a support with high porosity. The stabilizing effect was investigated in a series of glyceryl‐controlled‐pore glass (CPG) with varying specific surface areas (9.5–180 m2/g). The minimum addition of albumin, giving full stabilization, on the different supports correlated to a monolayer coverage of the surface, approximately 2–3 mg protein/m2. The effect of the additive was less pronounced when increasing amounts of enzyme were immobilized (5–40 mg enzyme/g Celite). The effect of the additives was studied using mandelonitrile lyase, but α‐chymotrypsin and lipase P were also shown to be stabilized. © 1993 John Wiley & Sons, Inc.
Kinetics of cell growth and heterologous glucoamylase production in recombinant Aspergillus nidulans
Authors: Lin Weng‐Long , Felberg Ross S. , De Bernardez Clark Eliana
Published: December 2004

In the work, a study of cell growth and the regulation of heterologous glucoamylase synthesis under the control of the positively regulated alcA promoter in a recombinant Aspergillus nidulans is presented. We found that similar growth rates were obtained for both the host and recombinant cells when either glucose or fructose was employed as sole carbon and energy source. Use of the potent inducer cyclopentanone in concentrations greater than 3 mM resulted n maximum glucoamylase concentration and maximum overall specific glucoamylase concentration over 80 h of batch cultivation. However, cyclopentanone concentrations in excess of 3 mM also showed an inhibitory effect on spore germination as well as fungal growth. In contrast, another inducer, threonine, had no negative effect on spore germination even when concentrations of up to 100 mM were used with either glucose or fructose as carbon source. Glucoamylase synthesis in the presence of glucose plus either inducer did not begin until glucose was totally depleted, suggesting strong catabolite repression. Similar results were obtained when fructose was employed, although low levels of glucoamylase were detected before fructose depletion, suggesting partial catabolite repression. The highest enzyme concentration (570 mg/L) and overall specific enzyme concentration (81 mg/g cell) were observed in batch culture when cyclopentanone was the inducer and fructose the primary carbon source. A maximum glucoamylase concentration of 1.1 g/L and an overall specific glucoamylase concentration of 167 mg/g cell were obtained in a bioreactor using cyclopentanone as the inducer and limited‐fructose feeding strategy, which nearly doubles the glucoamylase productivity from batch cultures. © 1993 John Wiley & Sons, Inc.
Production of cellobiose by enzymatic hydrolysis: Removal of β‐glucosidase from cellulase by affinity precipitation using chitosan
Authors: Homma Taira , Fujii Michihiro , Mori Jun‐ichi , Kawakami Tohru , Kuroda Kenji , Taniguchi Masayuki
Published: December 2004

Removal of β‐glucosidase (BG) from cellulase is essential to the enzymatic production of cellobiose from cellulose because of the high reactivity of BG with cellobiose to form glucose. Chitosan is a reversibly soluble‐insoluble polymer depending on pH, and it has an affinity with the other components, endo‐β‐1,4‐glucanase and cellobiohydrolase, or cellulase. The affinity precipitation technique using chitosan is an effective way to fractionate cellulase for the above purpose. Hydrolysis experiments of cellulose with the residual fractionated enzyme gave higher cellobiose contents in the soluble sugar products. © 1993 John Wiley & Sons, Inc.
Biofiltration of methanol vapor
Authors: Shareefdeen Zarook , Baltzis Basil C. , Oh Young‐Sook , Bartha Richard
Published: December 2004

Biofiltration of solvent and fuel vapors may offer a costeffective way to comply with increasingly strict air emission standards. An important step in the development of this technology is to derive and validate mathematical models of the biofiltration process for predictive and scaleup calculations. For the study of methanol vapor biofiltration, an 8‐membered bacterial consortium was obtained from methanol‐exposed soil. The bacteria were immobilized on solid support and packed into a 5‐cm‐diameter, 60‐cm‐high column provided with appropriate flowmeters and sampling ports. The solid support was prepared by mixing two volumes of peat with three volumes of perlite particles (i.e., peat–perlite volume ratio 2:3). Two series of experiments were performed. In the first, the inlet methanol concentration was kept constant while the superficial air velocity was varied from run to run. In the second series, the air flow rate (velocity) was kept constant while the inlet methanol concentration was varied. The unit proved effective in removing methanol at rates up to 112.8 g h−1 m−3 packing. A mathematical model has been derived and validated. The model described and predicted experimental results closely. Both experimental data and model predictions suggest that the methanol biofiltration process was limited by oxygen diffusion and methanol degradation kinetics. © 1993 John Wiley & Sons, Inc.
Catalytic and interfacial aspects of enzymatic polymer synthesis in reversed micellar systems
Authors: Rao A. Madhusudhan , John Vijay T. , Gonzalez Richard D. , Akkara Joseph A. , Kaplan David L.
Published: December 2004

The enzyme horseradish peroxidase, when encapsulated in reversed micelles, is capable of catalyzing the synthesis of phenolic and aromatic amine polymers. The synthesis of polyethylphenol is specifically considered in this article and is found to be extremely feasible in the micellar system. Polymer chain growth can be controlled to some degree by manipulating the ability of the solvent to sustain chain solubility; this is effectively done by adjusting the surfactant concentration. This results in a degree of control of polymer molecular weight. The synthesized polymer drops out of solution and can be easily recovered. © 1993 John Wiley & Sons, Inc.
Optimization of fed‐batch fermentors by iterative dynamic programming
Authors: Luus Rein
Published: December 2004

By using penalty functions to handle state constraints, iterative dynamic programming can be used in a straightforward manner for the optimization of fedbatch fermentors. No computational difficulties were encountered and better results are obtained than previously reported in the literature for a fed‐batch fermentor for biosynthesis of penicillin. © 1993 Johy Wiley & Sons, Inc.
Enhancement effect of polyethylene glycol on enzymatic synthesis of cephalexin
Authors: Hyun Chang K. , Choi Joon H. , Kim Jung H. , Ryu Dewey D. Y.
Published: December 2004

In an enzymatic synthesis of cephalexin (CEX) using an acylase from Xanthomonas citri, the effect of polyethylene glycol (PEG) on the synthetic reaction of 7‐amino‐3‐deacetoxycephalosporanic acid (7‐ADCA) and D‐alpha‐phenyl‐glycine methyl ester (PGM) to CEX was investigated. The addition of PEG (MW 300–20,000) increased the yield significantly. This yield enhancement effect tended to increase with the increasing molecular weight of PEG. Addition of PEG to the reaction system did not affect both the CEX and PGM hydrolytic reactions. The PEG added to the reaction medium used in these experiments did not depress the water activity significantly, and the product yield improvement could not be explained by the activity alone. The PEG stabilized the enzyme activity to some extent, but this stabilizing effect was only partially attributable to the yield enhancement of CEX. The enhancing effect of PEG on the synthetic yield increased with the increasing PEG molecular weight or the length of the poly(oxy‐1,2‐ethanediyl) chain, which increases the hydrophobicity of PEG. This finding consequently has led to the conclusion that the PEG structure renders the affinity between enzyme and 7‐ADCA, which is a hydrophobic substrate. The microenvironmental hydrophobicity of PEG and its interaction with the hydrophobic substrate was found to be the main reason for the improvement of the CEX yield. In fact, the Michaelis–Menten kinetic constant for 7‐ADCA, K7‐ADCA in the presence of PEG was smaller than that in the control system (without PEG addition). © 1993 John Wiley & Sons, Inc.
Cell retention culture with an internal filter module: Continuous ethanol fermentation
Authors: Chang Ho Nam , Lee Woo Gi , Kim Beom Soo
Published: December 2004

A new internal filter feedback system with a stainless steel filter was introduced and its application for continuous ethanol fermentation was investigated. The filter performance was highly influenced by agitation speed and yeast concentration. Retention coefficient with a filter of 2 μm pore size was found more than 97.5%, and the filter was suitable for yeast separation. Maximum yeast concentration was 157 g/L and the best operable cell concentration was between 90 and 150 g/L. Which was similar to that obtained in the external membrane cell recycle culture. The cell concentration in the fermentor was maintained by manipulation of dilution rate and bleed ratio with the growth rate. The internal filter feedback system was successfully operated for more than 10 days. This study shows that the internal filter feedback system with a stainless steel filter can be used high‐density cell culture and ethanol fermentation. Furthermore, it can be scaled up more easily than the external cell recycle system. © 1993 John Wiley & Sons, Inc.
A simulation model for the continuous production of acetoin and butanediol using Bacillus subtilis with integrated pervaporation separation
Authors: Dettwiler B. , Dunn I. J. , Heinzle E. , Prenosil J. E.
Published: December 2004

The potential for producing acetoin and butanediol with a Bacillus subtilis strain was investigated with continuous culture using molasses as carbon substrate. The steady‐state results were influenced by both oxygen and undetermined limiting compounds. Employing the known metabolic pathways, four overall stoichiometry relations were used with an energetic assumption on the energy requirements for biomass formation to establish a linear relations were used with an energetic assumption on the energy requirements for biomass formation to establish a linear relation between the overall rates, whose parameters were determined by linear regression. This provided a relationship for the product formation rate. The chemostat culture data were described with a growth kinetics model, which included limitation by molasses and oxygen as well as diauxic effects and product inhibition. The biokinetics model was combined with an experimentally verified model for the membrane Pervaporation. From this combined model were determined the influence of the membrane characteristics (enrichment factors and membrane area) and the dilution rate on the performance of the integrated process. Simulations revealed that an increase of the enrichment factor, possible by membrane improvement, would have counteracting influences, owing to decreased product inhibition but with lower biomass concentration. © 1993 Wiley & Sons, Inc.
Enhancement of cloned gene product synthesis via autoselection in recombinant Saccharomyces cerevisiae
Authors: Napp Scott J. , Da Silva Nancy A.
Published: December 2004

Saccharomyces cerevisiae autoselection strains with mutations in the ura3, fur1, and urid‐k genes have been obtained through a sequential isolation procedure. This autoselection system is an extension of one described by Loison et al. The mutations effectively block both the pyrimidine biosynthetic and salvage pathways and in combination are lethal to the host. Therefore, a plasmidencoded URA3 gene is essential for cell viability regardless of the growth conditions, and complex (traditionally nonselective) media can be employed without the risk of plasmid loss. The effects of medium enrichment on growth and cloned gene product synthesis were examined in batch culture for two autoselection strains. The plasmid gene product β‐galactosidase was under the control of the yeast GAL1 promoter, and two methods of induction were employed; one strain was induced via temperature shift while the other was induced by galactose addition. Three nutrient media were investigated: a lean selective medium (SD), a richer semidefined medium (SDC), and a rich complex medium (YPD). The results demonstrated the improvements in cloned gene productivity possible when the growth medium is enriched, with up to 10‐fold increases in β‐galactosidase productivity observed. Plasmid instability and mutation reversion were not problems for the autoselection strains, even in uracil‐containing medium. Short‐term plasmid stabilities were approximately 90% in all three media tested. During continuous culture of the autoselection temperature‐sensitive strain, long‐term plasmid stability was excellent and β‐galactosidase expression remained high after more than 25 residence times under inducing conditions. In contrast, both β‐galactosidase specific activity and plasmid stability decreased linearly with time for an analogous nonautoselection strain. The introduced fur1 and uridk mutations were very stable; after more than 50 generations of growth in complex medium, stability values of 99–100% were measured. © 1993 Wiley & Sons, Inc.
Fluid‐mechanical forces in agitated bioreactors reduce the CD13 and CD33 surface protein content of HL60 cells
Authors: Lakhotia Sanjay , Bauer Kenneth D. , Papoutsakis Eleftherios T.
Published: December 2004

Flow cytometry was used to examine the effect of hydrodynamic forces in surface aerated stirred tank bioreactors on the quantity of CD13 and CD33 surface proteins of Hl60 (human promyelocytic leukemia) cells. A step increase in agitation of the 2‐L bioreactors from 80 to 400 rpm reduced the apparent growth rate and the average CD13 and CD33 content per HL60 cell. The effects on the two surface proteins were observed within 30‐60 min following the increase in the agitation and preceded observed effects on cell growth by at least 10 h. Upon reduction of the agitation rate back to 80 rpm, the CD13 and CD33 content recovered (in ca. 10 h) for CD13 and ca. 29h for (CD33) to the levels of the control culture whose agitation rate was maintained at 80rpm. The CD13 and CD33 cell content was reduced even at agitation rates (270 rpm) that did not affect cell proliferation. Pluronic F68 (a commonly used shear protectant) had a protective effect on the CD33 content per cell of cultures subjected to hydrodynamic injury but no effect on the CD13 cell content. Possible bioprocessing and physiological implications of these findings are discussed © 1993 Wiley & Sons, Inc.