bluuebunny/crossref_metadata_2025 huggingface dataset:
Fed‐batch culture automated by uses of continuously measured cell concentration and culture volume
Authors: Yamane Tsueno , Hibino Wataru , Ishihara Kazuto , Kadotani Yasuyuki , Kominami Minoru
Published: December 2004

An automated control method of fed‐batch culture in which the nutrient feed rate was determined from continuously measured cell concentration and culture broth volume was developed. Theoretical background was elucidated, from which it was found that the method is unique in that it controls specific substrate consumption rate of the microorganism. The method was experimentally applied to the fed‐batch cultures of recombinant Escherichia coli HB101. It was observed that the specific substrate feed rate affects not only the specific growth rate but also the growth yield. If some conditions are satisfied, this type of automated fedbatch culture can be applied widely to any microbial systems and seems especially useful when the culture medium is composed of natural complex nutrient(s) because their concentrations are very difficult to detect and control.
Extractive lactic acid fermentation using ion‐exchange resin
Authors: Srivastava Aradhana , Roychoudhury Pradip K. , Sahai Vikram
Published: December 2004

Lactic acid fermentation is an end‐product‐inhibited reaction. The restriction imposed by lactic acid on its fermentation can be avoided by extractive fermentation techniques. Studies were performed by attaching an ion‐exchange resin packed column with a 2‐L fermentor for separation of lactic acid. The fermentation, in a conventional batch mode, resulted in a lactic acid yield of 0.828 g · g−1 and a lactic acid productivity of 0.313 g · L−1 · h−1. However, these could be further enhanced to 0.929 g · g−1 and 1.665 g · L−1 · h−1 by extractive fermentation techniques. The effect of temperature on extractive fermentation was remarkable and has been included in this work.
Kinetics of enzymatic degradation of cyanide
Authors: Basheer S. , Kut Ö. M. , Prenosil J. E. , Bourne J. R.
Published: December 2004

CYANIDASE@ is a new enzyme preparation capable of degrading cyanide in industrial wastewaters to ammonia and formate in an apparently one‐step reaction, down to very low concentrations. This enzyme has both a high selectivity and affinity toward cyanide. A granular form of the biocatalyst was used in a recirculation fixed bed reactor in order to characterize the new biocatalyst with respect to pH, ionic strength, common ions normally present in wastewaters, mass transfer effects, and temperature. Long term stability was investigated. The kinetics of the enzymatic degradation of cyanide were studied in a batch reactor using the powdered immobilized enzyme preparation and modeled using a simple Michaelis‐Menten equation.
Random mutagenesis to enhance the activity of subtilisin in organic solvents: Characterization of Q103R subtilisin E
Authors: Economou Chariklia , Chen Keqin , Arnold Frances H.
Published: December 2004

Q103R subtilisin E was isolated following random mutagenesis and screening for improved activity in the presence of dimethylformamide (DMF). Our goal is to identify the mechanism(s) by which amino acid substitutions can enhance enzyme activity in polar organic solvents. A quantitative framework for comparing substrate binding and catalytic activities of mutant and wild‐type enzymes in the presence and absence of DMF is outlined. Kinetic experiments performed at high salt concentration (1M KCl) reveal that the mechanism behind the Q103R variant's enhanced activity toward succinyl‐Ala‐Ala‐Pro‐Phe‐p‐nitroanilide is both electrostatic and nonelectrostatic in origin. Favorable electrostatic interactions between the negatively charged succinyl group of the substrate and the positive charge on Arg 103 are responsible for tighter substrate binding. This conclusion is supported by kinetic experiments performed on the related substrate Ala‐Ala‐Pro‐Phe‐p‐nitroanilide and the hydrolysis kinetics of the Q103E, Q103K, and Q103S variants constructed by site‐directed mutagenesis. These results highlight the importance of the choice of the substrate used to screen for improvements in catalytic activity.
Enzyme‐based strategy for toxic waste treatment and waste minimization
Authors: Smith Jeffrey M. , Payne Gregory F. , Lumpkin Janice A. , Karns Jeffrey S.
Published: December 2004

The increasing amounts of pesticides used throughout the world, as well as the increasingly stringent governmental regulations concerning waste disposal, mandates improved techniques of waste disposal and minimization. In this article, parathion hydrolase, an enzyme with proven effectiveness at hydrolyzing organophosphates, was used to treat a cattle dipping liquid containing the pesticide, coumaphos, which is used to kill a disease‐causing tick. Waste is generated from this process when a toxic dechlorination product of coumaphos, potasan, accumulates to concentrations hazardous to the cattle. This pesticide system was used as a model to demonstrate how enzyme technology can be applied to waste treatment and minimization. Kinetic experiments showed that the hydrolysis of the two organophosphate substrates can be modeled as first‐order reactions with identical rate constants. It was further shown that the enzyme is capable of hydrolyzing only dissolved substrates. Because of the eightfold greater solubility of potasan than coumaphos (16.9 vs. 2.2 μmol/L), it was possible to utilize the enzyme to hydrolyze potasan selectively. Thus, by limiting the amount of enzyme, it is possible to remove potasan selectively to extend the lifetime of the cattle dipping liquid, thereby reducing the amount of waste generated. Based upon experimental results, a mathematical model describing the system was developed and verified. The mathematical model was then used to simulate the ability of the enzyme to hydrolyze the total amount of organophosphates, and to degrade selectively all of the toxic potasan without a significant loss of coumaphos.
Synthesis and lysis of formate by immobilized cells of Escherichia coli
Authors: Nandi R. , Bhattacharyya P. K. , Bhaduri A. N. , Sengupta S.
Published: December 2004

Formate hydrogenlyase (FHL) activity was induced in a strain of Escherichia coli S13 during anaerobic growth in yeast extract‐tryptone medium containing 100 mM formate. The cells obtained at the optimum growth phase were immobilized in 2.5% (w/v) agar gel when 50–60% of the whole cell FHL activity was retained. The immobilized FHL system had good storage stability and recycling efficiency. In the lysis of formate, an increase of formate concentration to 1.18M increased QH2 (initial) value of the immobilized cell, and subsequently cells, hydrogen evolution, in general, ceased after 6 to 8 of incubation, resulting in incomplete lysis of formate. Presence of small amount of glucose (28 mM) was more or less quantitatively lysed with concomitant disappearence of glucose from the medium. Synthesis of formate from hydrogen and bicarbonate solution by the immobilized cells was also characterized. Presence of glucose (10 mM) in 50 mM bicarbonate solution stimulated formate synthesis by immobilized cells. The pH optimum range, Km, and specific activity of the immobilized cells for the lysis of formate were 6.8–7.2 0.4M, and 66 mL/g cell‐h, respectively. The cells could fix hydrogen to the extent of 24.4% (w/w) of its own wet cell mass in a 72‐h reaction cycle. Potentiality of the immobilized FHL system for biotechnological exploitation was discussed.
Determination of the interparticular effective diffusion coefficient for CO2 and O2 in solid state fermentation
Authors: Auria Richard , Palacios Jorge , Revah Sergio
Published: December 2004

A simple experimental diffusion controlled fermentor (DCF), coupled with the use of a mathematical model based on mass balance, is proposed to measure the variation of the gas (CO2 and O2) diffusion coefficients in solid state fermentation. The DCF was packed with an ion‐exchange resin impregnated with a nutritive medium and inoculated with Aspergillus niger. The growth conditions in the DCF were very similar to those found in equipment operated with convective oxygen supply. The diffusion coefficient was shown to be very dependent on the biomass concentration within the solid state fermentor, and attained values of less than 5% of the molecular diffusion in air when the biomass in the fermentor reached 27 mg dry/g dry support.
Cultivation characteristics of immobilized Aspergillus oryzae for kojic acid production
Authors: Kwak Moo Young , Rhee Joon Shick
Published: December 2004

Aspergillus oryzae in situ grown from spores entrapped in calcium alginate gel beads was used for the production of kojic acid. The immobilized cells in flask cultures produced kojic acid in a linear proportion while maintaining the stable metabolic activity for a prolonged production period. Kojic acid was accumulated up to a high concentration of 83 g/L, at which the kojic acid began to crystallize, and, thus, the culture had to be replaced with fresh media for the next batch culture. The overall productivities of two consecutive cultivations were higher than that of free mycelial fermentation. However, the production rate of kojic acid by the immobilized cells was suddenly decreased with the appearance of central cavernae inside the immobilized gel beads after 12 days of the third batch cultivation.
Effect of applied potentials on the activity and growth of Thiobacillus ferrooxidans
Authors: Natarajan K. A.
Published: December 2004

The effect of applied DC potentials both in the positive and negative range, on the activity and growth of Thiobacillus ferrooxidans, is discussed. In general, application of positive potentials up to +1000 mV in an acid bioleaching medium was found to be detrimental to bacterial activity, while the impression of negative potentials enhanced both their activity and growth through electrochemical regeneration of ferrous ions and an increase in the biomass. Ferrous–ferric ratios in a bioleaching medium could be monitored through Eh measurements.

Among the base sulfide minerals such as pyrite, chalcopyrite, and sphalerite, sphalerite could be selectively bioleached if an impressed potential of −500 mV (SCE) could be maintained in the leaching medium. Electrochemical bioleaching tests carried out under an applied potential of −500 mV with sphalerite in the presence and absence of noble minerals such as pyrite and chalcopyrite indicated enhanced zinc dissolution with negligible copper and iron in solution. Probable mechanisms and advantages of the electrochemical bioleaching process developed in the laboratory are outlined.
Effects of glucose on the production of recombinant protein C in mammalian cell culture
Authors: Sugiura Takeyuki
Published: December 2004

Effects of glucose on a cultured Chinese hamster ovary cell line producing recombinant human protein C were investigated. After the recombinant cells reached confluency, they were maintained in the medium containing 10% serum and different levels of glucose in either batch or daily‐exchange mode. High concentrations of glucose to the cultures yielded higher cell densities. Daily exchanges of media produced higher cell densities than the corresponding batch culture. Total protein C production per cell decreased with time in batch culture, in accordance with the declined glucose metabolism. Supplementation of the media with high levels of glucose diminished both the expression and γ‐carboxylation activities of the recombinant cells. Production of protein C persisted in daily‐exchange culture, resulting in a constant production rate of protein C. In this case again, glucose reduced the specific productivity of recombinant protein C. An apparent glucose inhibition constant was determined to be 0.11 mg/mL by Dixon plots. The ability to γ‐carboxylate recombinant protein C was also impaired at the highest level of glucose. From these results, a strategy to maximize recombinant protein C productivity is discussed.
Analysis and quantification of a mixed exo‐acting and endo‐acting polysaccharide depolymerization system
Authors: Dean Sheldon W. , Rollings James E.
Published: December 2004

A new mathematical model has been proposed based on a model presented by Suga, van Dedem, and Moo‐Young.10 The model requires a separate differential equation for each polymeric species (differentiated by degree of polymerization) in the reaction mixture. The main contribution of this model is the incorporation of experimental molecular weight distributions as the initial conditions. These molecular weight distributional as the initial conditions were obtained using modern analytical equipment previouly unknown for this application. The equipment, SEC/LALLS, measures relative concentrations of specific molecular weight species along with the corresponding molecular weights, thus yielding (through some mathematical manipulation) the absolute concentration of each molecular weight species. The concentration at each molecular weight can then be incorporated as the initial condition for that equation. Theoretically, the system of differential equations can be solved to give a more realistic time course of reaction.

Synergism between endo‐acting and exo‐acting enzymes was examined theoretically using the mathematical model. Through model predictions, it was found that synergy is based on two fundamental parameters: (1) each enzyme's activity relative to the sum of enzyme activities and, (2) overall substrate concentration relative to the exo‐acting enzyme's Michaeiis kinetic constant Km. Theoretically, synergism increases as a function of reaction time. Intermediate endo fractions (ratio of endo‐acting enzyme activity to the sum of endo‐acting and exo‐acting enzyme activity) from 0.3 to 0.7 exhibit the most synergism. Values of k[log(Km, exo/S0)] above about zero also exhibits the most synergism.

An examination of experimental data obtained both by SEC/LALLS and by reducing sugar measurements shows that the model is inadequate for successfully predicting quantities associated with the substrate during reaction. This is especially true for synergism predictions. At short reaction times, the model predicts the data fairly well, but at longer times the predictions are inconsistent with experimental data. These inconsistencies may be due to complicating phenomena such as enzyme inhibitions.
Alkaloid production by plant cell suspension cultures of Holarrhena antidysenterica: I. Effect of major nutrients
Authors: Panda A. K. , Mishra Saroj , Bisaria V. S.
Published: December 2004

The effect of major nutrients on growth and alkaloid production by plant cell culture of Holarrhena antidysenterica was studied with a view to increasing the yield of the alkaloid conessine, a therapeutic drug used for treatment of dysentery and helminthic disorders. The studies resulted in development of a modified Murashige and Skoog (MS) medium that contained 60 mM total nitrogen with a NH4+‐to‐NO3− ratio of 5:1, 0.25 mM phosphate, and 40 g/L sucrose. The growth regulators 2,4‐dichlorophenoxy acetic acid (2,4‐D) and kinetin (Kn) were also found to affect the synthesis of alkaloid. Using an optimal level of inoculum (3 g/L), the modified medium resulted in alkaloid synthesis of 0.66 g/100 g dry cell weight, which represented a 4.25‐fold increase over that obtained in standard MS medium.
Alkaloid production by plant cell cultures of Holarrhena antidysenterica: II. Effect of precursor feeding and cultivation in stirred tank bioreactor
Authors: Panda A. K. , Bisaria V. S. , Mishra Saroj
Published: December 2004

Precursor feeding strategy for increasing the yield of conessine, a steroidal alkaloid of Holarrhena antidysenterica, was established in cell suspension culture. A total of 50 mg/L added cholesterol was converted into 43 mg/L of alkaloid, 90% of which constituted the conessine. By applying the precursor feeding policy to the cell suspension culture in modified Murashige and Skoog (MS) medium, a total of 143 mg/L of alkaloid was produced in 8 days. In this way the alkaloid content of the cells was increased more than six times compared to that obtained in the standard MS medium. The steps leading to biotransformation of cholesterol into alkaloids were unaffected by phosphate. The shake flask data were successfully transferred to a bench scale 6‐L stirred tank bioreactor in which the specific biosynthetic rate of alkaloid production was 110 mg/100 g dry cell weight per day, about 160 times higher than that of whole plant.
Modeling and experimental validation of carbon dioxide evolution in alkalophilic cultures
Authors: Noorman H. J. , Luijkx G. C. A. , Luyben K. Ch. A. M. , Heijnen J. J.
Published: December 2004

The chemical reactions involving carbon dioxide in mineral culture media are considered. A mathematic model is set up, based on published data, which is valid at pH values below 9, and in which the nonideality of the solution is taken into account. The crucial parameter is the constant expressing the equilibrium between carbon dioxide and bicarbonate, K1.

The reactions were studied in three different aqueous solutions: water, mineral salt medium, and a suspension with nongrowing bacterial cells. For each situation, three methods were compared for the determination of the bicarbonate concentration in the solution: equilibrium state total carbon analysis, dynamic monitoring of the rate of acid or alkali addition, and dynamic measurement of the carbon dioxide gas phase mole fraction.

In a batch‐stirred tank reactor, the equilibrium constant K1 agreed with the published value, and the three bicarbonate analysis methods give the same results. If the nonideality is not taken into account, the result significantly differed from the published value and is likely to be incorrect.

A real alkalophilic process, using Acinetobacter calcoaceticus in a continuous stirred tank reactor at steady state, also gave results that are in accord with the literature. However, the results do not allow validation of the equation expressing the nonideality.

The steady state in the batch system and in continuous culture can be well described with the mathematical model. However, in the transient state there are some unexplained differences between simulation and measurement.
Inner mitochondrial membranes bound to concanavalin A–sepharose display succinate dehydrogenase, ATPase, and cytochrome oxidase activity
Authors: Strasser Reto J. , Millán Lourdes , Darszon Alberto
Published: December 2004

A fraction (15–20% of the total protein) of a preparation of bovine submitochondrial particles (SMPs) binds to concanavalin A–sepharose. The bound membranes displayed succinate dehydrogenase, cytochrome oxidase, and ATPase activity, which, as in SMPs, were inhibited by malonate, cyanide, and oligomycin, respectively. These results indicate that the bound membranes are inner mitochondrial membranes and that they contain a glycoprotein which was recognized by concanavalin A. It was possible to repeatedly perform the three enzyme assays, one after the other, in the same gel with the bound membranes. Long‐term stability tests (22 days) showed that cytochrome oxidase was much more stable in the membranes bound to the gel than in SMPs, while the ATPase activity decayed at a similar rate in the two conditions. Thus, inner mitochondrial membranes bound to ConA–Sepharose appear to be a potentially interesting model for the study of immobilized multienzymatic complexes.
Solubilization and regeneration of Vitreoscilla hemoglobin isolated from protein inclusion bodies
Authors: Hart Roger A. , Bailey James E.
Published: December 2004

Vitreoscilla hemoglobin (VHb), a homodimeric protein containing two heme groups in its native state, was used as a model to investigate inclusion body approtein solubilization, prosthetic group incorporation, and reactivation. High‐level expression in recombinant Escherichia coli results in accumulation of a substantial portion of heme‐free VHb in inclusion bodies. VHb can be solubilized from these inclusion bodies by relatively low concentrations of urea with the dissolution midpoint at approximately 3.2M urea. Dissolution in the presence of stoichiometric heme shifts the dissolution midpoint to approximately 4.5M urea without influencing the dissolution properties of contaminant proteins, suggesting the effect is specific for VHb. Denaturation of apoVHb and holoVHb obtained from purified native VHb has midpoints of 2.9M and 5.1M urea, respectively. VHb solubilized from inclusion bodies with urea at concentrations from 0 to 3.5M urea can be regenerated by heme addition without dilution of urea to yield active holoVHb. The fraction of solubilized VHb reconstituted upon heme addition is maximum at around 30% when solubilization and reconstitution is conducted in less than 1M urea. At these low urea concentrations, approximately 5% of inclusion body VHb is solubilized. These results show the utility of prosthetic group addition to reconstitute holoVHb in the presence of urea. Also, these findings suggest that some inclusion body protein has partially folded conformation and that a fractional dissolution and refolding process may be advantageous.
Changes of lipase‐catalyzed lipolytic rates in a batch reactor
Authors: Wang Y. J. , Wang F. F. , Sheu J. Y. , Tsai Y. C. , Shaw J. F.
Published: December 2004

A dramatic change of the reaction rate was observed for the lipase‐catalyzed hyrolysis of tributyrin in a batch reactor. Immediately after the addition of the enzyme, the lipolysis rate increased continuously until a maximal reaction rate was reached. The duration of the induction was mainly controlled by the bulk enzyme concentration and the reactor stirring speed. The reaction rate dropped sharply after reaching its maximal value. The lipolysis decayed at a rate of about 0.012 min−1, and was not affected by changes of the stirring speed. This decay was attributed to the fast deactivation of the surface‐adsorbed lipase, and possibly to the extremely slow desorption of the inactivated species. For reaction time longer than 120 minutes, the lipolysis decreased at a much slower rate. Several mechanisms for the decay of the lipolysis rate were discussed.
A fluorescence‐based fiber optic biosensor for the flow‐injection analysis of penicillin
Authors: Xie Xiangfang , Suleiman Ahmad A. , Guilbault George G.
Published: December 2004

A penicillin fiber optic sensor is described. The sensor is based on co‐immobilization of a pH indicator, fluorescein isothiocyanate (FITC), and penicillinase on a preactivated biodyne B membrane attached to the end of a bifurcated optical fiber. The characteristics of the sensor are investigated in conjunction with a flow injection analysis system. The proposed sensor is reversible and responds to penicillin in the concentration range of 1 × 10−4 to 5 × 10−2 mol/L. The application of this sensor to penicillin analysis in some pharmaceutical samples is demonstrated.
Fermentation of sweet whey by ethanologenic Escherichia coli
Authors: Guimaraes Walter V. , Dudey Gregory L. , Ingram L. O.
Published: December 2004

Whey, an abundant byproduct of the dairy industry, contains large amounts of protein and lactose which could be used for fuel ethanol production. We have investigated a new organism as a candidate for such fermentations: recombinant Escherichia coli containing the genes encoding the ethanol pathway from Zymomonas mobilis. The highest level of ethanol achieved, 68 g/L, was produced after 108 hours in Luria broth containing 140 g lactose/L. Fermentations of lower lactose concentrations were completed more rapidly with approximately 88% of theoretical yields. Reconstituted sweet whey (60 g lactose/L)was fermented more slowly than lactose in Luria broth requiring 144 hours to produce 26 g ethanol/L. Supplementing sweet whey with a trace metal mix and ammonium sulfate reduced the required fermentation time to 72 hours and increased final ethanol concentration (28 g ethanol/L). By adding proteinases during fermentation, the requirement for ammonia was completely eliminated, and the rate of fermentation further improved (30 g ethanol/L after 48 hours). This latter incresed in rate of ethanol production and ethanol yield are presumed to result from incorporation of amino acids released by hydrolysis of whey proteins. The fermentation of sweet whey by ethanologenic E. coil reduced the nonvolatile residue by approximately 70%. This should reduce biological oxygen demand and reduce the cost of waste treatment. Whey supplemented with trace metals and small amounts of proteinase may represent an economically attractive feedstock for the production of ethanol and other useful chemicals.
A comparison of neural networks and partial least squares for deconvoluting fluorescence spectra
Authors: McAvoy Thomas J. , Su Hong Te , Wang Nam S. , He Ming , Horvath John , Semerjian Hratch
Published: December 2004

This article compares backpropagation neural networks (BNN) with partial least squares (PLS) techniques in terms of their ability to deconvolute fluorescence spectra. Both actual experimental and simulated spectral data are studied for 2 binary systems. These systems consist of mixtures of tryptophan and tyrosine, and NADH and tryptophan over a total concentration range of 10−7 to 10−4 M. It is shown that BNN is superior to PLS for both systems.
Mechanisms of protein solubilization in reverse micelles
Authors: Matzke S. F. , Creagh A. L. , Haynes C. A. , Prausnitz J. M. , Blanch H. W.
Published: December 2004

Solubilization properties of α‐chymotrypsin and alcohol dehydrogenase (LADH) in reverse micelles are reported for three different solubilization techniques. The solubilization properties for these two proteins depend on the method used for protein addition. The addition of a dry protein powder to a reverse‐micelle‐containing organic phase does not appreciably solubilize the protein until the diameter of the reverse micelle is similar to that of the protein. However, when an aqueous protein solution is injected an organic phase, protein solubilization is not strongly dependent on micelle size. For chymotrypsin, multiple protein occupancy occurs at large micelle size, with as many as 11 chymotrypsin molecules solubilized in one reverse micelle. The solubilization of chymotrypsin using a phase‐transter technique with a positively charged surfactant follows the expected traned based on protein‐surfactant electrostatic interactions. When a negatively charged sufactants is used for phase transfer, at low pH the solubilization data do not fit this electrostatic interaction mechanism. In this case, proteinsurfactant aggregation may be occurring at the aqueousorganic interface.
Use of microbial electrodes for observation of microbiological nitrogen elimination process
Authors: Hikuma Motohiko , Matsuoka Hiroshi , Kawarai Masahiro , Karube Isao
Published: December 2004

Microbial electrodes for the determination of ammonia and the estimation of biochemical oxygen demand (BOD) were applied to the nitrogen elimination process. The dimensions of the nitrification and the denitrification vessels were 170 and 70 L, respectively. The wastewater used for the experiment was obtained from a fermentation factory and adjusted to 470–530 mg/L of total Kjeldahl nitrogen and 1700–3000 mg/L of BOD. The ammonia electrode was assembled with a membrane containing nitrifying bacteria and an oxygen probe. The BOD electrode was similarly constructed, except it used a membrane containing the yeast, Trichosporon cutaneum. A flow system was employed for the automatic measurement of samples every 30 min. The nitrification and denitrification rates of the activated sludge were measured to investigate the optimum conditions and evaluate the capacity of the plant. The various data obtained by the microbial electrodes allowed us to inspect the situation of the plant and estimate control paramerers such as nitrogen and BOD loadings. The average removals of ammonia nitrogen and total Kjeldahl nitrogen were 96% and 89%, respectively, during the experiment for period of 2 weeks.
Photosynthetic activity of cyanobacteria in water‐in‐oil microemulsions
Authors: Famiglietti M. , Hochköppler A. , Wehrli E. , Luisi P. L.
Published: December 2004

The solubilization and the photosynthetic activity of cyanobacteria (Anabaena variabilis) in water‐in‐oil microemulsions consisting of (Tween85/Span80)/hexadecane/water is investigated. Transparent and stable solutions containing up to 108 cells/mL could be obtained. The physical state and stability of the cells in the microemulsion, as evidenced from optical and electron microscopy, is dependent upon the physical parameters of the system, and in particular on the hydrophylic‐lypophilic balance (HLB) of the surfactant system. Conditions could be found, under which the cells in the microemulsion system display photosynthetic activity This was judged by measuring polarographically the oxygen evolution and by studying the photosynthetic activity in the presence of specific inhinbitors.
Functional immobilization of lipase eliminating lipolysis product inhibition
Authors: Kosugi Yoshitsugu , Suzuki Hideo
Published: December 2004

Lipase from Pseudomonas fluorescens biotype I was immobilized on macroporous anion exchange resin using glutaraldehyde to enhance the adsorption. The immobilization method was selected, because it provided the highest extent of hydrolysis of beef tallow at high substrate concentrations. The immobilized lipase was not substantially inhibited by oleic acid or sodium oleate, but the soluble lipase was strongly inhibited by both substances. The optimum pH of lipolysis was pH 4 for the immobilized lipase and pH 6 for the soluble one. These results indicate that the microenvironment created around the lipase molecule by immobilization eliminates product inhibition. In addition, the immobilization on the support enhances the stability of the lipase against chemical denuaturation. © 1992 John Wiley & Sons, Inc.
Effect of oxygen on steady‐state product distribution in Bacillus polymyxa fermentations
Authors: Mankad T. , Nauman E. B.
Published: December 2004

Bacillus polymyxa ferments glucose to 1‐2,3 butanediol, acetoin, ethanol, acetic acid, lactic acid, and formic acid. This research investigates product formation as a function of oxygen availability. A predictive model that simulates product distribution at known oxygen transfer rates is developed on the hypothesis that, in an energy‐limited environment, B. polymyxa utilizes glucose and oxygen in the most efficient manner. The efficiency of utilization of glucose and oxygen is measured in terms of the ATP yields of each oxidative pathway. The identity of the products constituting the profile at the given oxygen transfer rate is determined by comparing the ATP production and consumption rates. While the ATP generated is calculated from a knowledge of the oxygen transfer rate and ATP yields of the oxidative pathways, the ATP consumption is estimated by the Pirt expression in terms of growth‐ and nongrowth‐associated components. The product formation rates are obtained by solving ATP and NAD balance equations. They equate the production and consumption rates of these intermediates and are derived from the pseudo‐steady‐state hypothesis. The model is applied to continuous culture systems that are both open and closed with respect to biomass. At a given oxygen transfer rate, dilution rate, and inlet glucose concentration, the model predicts steady‐state concentrations of two dominant fermentation endproducts with the help of four parameters that can be determined from independent experiments. In contrast with earlier approaches, the experimental studies are carried out in continuous culture. Product profiles are obtained at various oxygen transfer rates, fer rates, inlet glucose concentrations, and dilution rates. The effect of pH on the relative distribution of products is also demonstrated. Results indicate that the model is fairly successful in predicting product profiles as a function of oxygen availability. © 1992 John Wiley & Sons, Inc.
Growth kinetics of the photosynthetic bacterium Chlorobium thiosulfatophilum in a fed‐batch reactor
Authors: Kim Byung Woo , Chang Ho Nam , Kim In Kyu , Lee Kang Suk
Published: December 2004

Hydrogen sulfide dissolved in water can be converted to elementary sulfur or sulfate by the photosynthetic bacterium Chlorobium thiosulfatophilum. Substrate inhibition occurred at sulfide concentrations above 5.7 mM. Light inhibition was found at average light intensities of 40,000 lux in a sulfide concentration of 5 mM, where no substrate inhibition occurred. Light intensity, the most important growth parameter, was attenuated through both scattering by sulfur particles and absorption by the cells. Average cell and sulfur particle sizes were 1.1 and 9.4 μm, respectively. Cells contributed 10 times as much to the turbidity as sulfur particles of the same weight concentration. The light attenuation factor was mathematically modeled, considering both the absorption and scattering effects based on the Beer‐Lambert law and the Rayleigh theory, which were introduced to the cell growth model. Optimal operational conditions relating feed rate vs. light intensity were obtained to suppress the accumulation of sulfate and sulfide and save light energy for 2‐ and 4‐L fed‐batch reactors. Light intensity should be greater for the same performance (H2S removal rate/unit cell concentration) in larger reactors due to the scaleup effect on light transmission. Knowledge of appropriate growth kinetics in photosynthetic fed‐batch reactors was essential to increase feed rate and light intensity and therefore cell growth. A mathematical model was developed that describes the cell growth by considering the light attenuation factor due to scattering and absorption and the crowding effect of the cells. This model was in good agreement with the experimental results. © 1992 John Wiley & Sons, Inc.
Effect of hydrogen sulfide on growth of sulfate reducing bacteria
Authors: Reis M. A. M. , Almeida J. S. , Lemos P. C. , Carrondo M. J. T.
Published: December 2004

A culture of sulfate reducing bacteria (SRB) growing on lactate and sulfate was incubated at different pH values in the range of 5.8–7.0. The effect of pH on growth rate was determined in this pH range; the highest growth rate was observed at pH 6.7. Hydrogen sulfide produced from sulfate reduction was found to have a direct and reversible toxicity effect on the SRB. A hydrogen sulfide Concentration of 547 mg/L (16.1 mM) completely inhibited the culture growth. Comparison between acetic acid and hydrogen sulfide inhibition is presented and the concomitant inhibition kinetics are mathematically described. © 1992 John Wiley & Sons, Inc.
Iridium oxide pH microelectrode
Authors: VanHoudt Paulus , Lewandowski Zbigniew , Little Brenda
Published: December 2004

The manufacture, calibration, and signal conditioning during construction of an iridium/iridium oxide pH microsensor is described. The microsensor was designed to be used extracellularly, primarily in biofilm research. The sensing tip diameters were typically in the range of 3–15 μm. The iridium oxide was formed by potential cycling in dilute sulfuric acid. A pH profule across a denitrifying biofilm was measured as an example of an application. The higher Nernstian slope (70–80 mV/pH for fresh electrodes), increased rigidity, and restriction of the sensing tip to the outermost end of the electrode are features which make the iridium/iridium oxide pH microelectrode superior to a glass microelectrode. © 1992 John Wiley & Sons, Inc.
Investigation of production of dextran and dextransucrase by Leuconostoc mesenteroides immobilized within porous stainless steel
Authors: El‐Sayed Abdel‐Halim M. M. , Abdul‐Wahid Kamsani , Coughlin Robert W.
Published: December 2004

Cells of Leuconostoc mesenteroides were immobilized within porus, stainless‐steel (SS) supports and used for dextransucrase (DS) and dextran production. The pore size of the support significantly affected the dextran yields, which were greatest with average pore sizes of 2‐5 μm. All immobilized‐cell biocatalysts in porous stainless steel produced higher yields than free cells, with the exception of cells confined in submicrometer pores (0.5 μm). Coating supports of larger pore size (40 and 100 μm) with calcium alginate enhanced the cell‐loading capacity of the supports and increased dextran and fructose yields in the cell‐free broth. Controlled, fed‐batch, DS production (activation), as a step preliminary to dextran production, significantly improved the subsequent dextran and fructose yields and shortened the time required to attain the maximum such yields. Scanning electron microscopy (SEM) of immobilized L. mesenteroides in stainless steel shows an irregular pattern of the microorganism inside the pores of the solid supports. Coating the porous solid supports with a cell‐free calcium alginate layer led to an increase in the cell density inside the support. Cell growth inside the coated, porous stainless steel had no distinct growth form. © 1992 John Wiley & Sons, Inc.
Kinetics of methanogenic degradation of phenol by activated‐carbon‐supported and granular biomass
Authors: Craik S. A. , Fedorak P. M. , Hrudey S. E. , Gray M. R.
Published: December 2004

A continuous‐feed recycle bioreactor was used to study the kinetics of methanogenic degradation of phenol at 35°C by bacteria supported on a bed of granular activated carbon (GAC). At dilution rates well above the growth rate of the culture, the cells not only populated the GAC, but also formed a layer of granular biomass. This layer was stabilized by the presence of the GAC, and accounted for over half of the phenol‐degrading activity in the bioreactor. The specific phenol degradation rates for GAC‐attached biomass, suspended biomass, and granular biomass were all in the range 0.15 to 0.22 mg phenol/mg volatile solids per day as measured under pseudo‐steady‐state conditions. © 1992 John Wiley & Sons, Inc.
Mammalian cell retention in a spinfilter perfusion bioreactor
Authors: Yabannavar V. M. , Singh V. , Connelly N. V.
Published: December 2004

A spinning cylindrical filter is often used to retain mammalian cells in a continuous perfusion bioreactor. This device, known as a spinfilter, has typically been with pore size smaller than the cell particles (single cells or aggregates) in order to achieve cell separation. For single cells in suspension, such an operation cannot be sustained over a long period of time because of clogging of the filter surface. Recently, screens with openings larger than the average cell size have been used to reduce the incidence of clogging. In this article, we have investigated how the screen size affects cell retention. We also showed why it is necessary to optimize the rotational speed of the spinfilter in order to achieve cell retention and reduce screen clogging. Effects of bulk mixing and perfusion rate on screen fouling cell retention, and cell washout were also investigated. © 1992 John Wiley & Sons, Inc.
Maximization of lipid production by Rhodotorula gracilis CFR‐1 using response surface methodology
Authors: Prapulla S. G. , Jacob Z. , Chand Nagin , Rajalakshmi D. , Karanth N. G.
Published: December 2004

In this study, an attempt has been made to maximize the lipid production using response surface methodology (RMS). Levels of nitrogen, carbon, and inoculum were chosen as factors. The results indicated that inoculum level was very important in lipid production, followed by carbon and nitrogen levels. At higher levels of inoculum, the strain was found to be more tolerant to higher concentrations of sugar, and significantly increased lipid production was noticed. Through the fitted models of second order, as per RSM, carbon at 10.24%, nitrogen at 0.37 g/L, and inoculum at the 20% level resulted in maximum biosynthesis of lipids. © 1992 John Wiley & Sons, Inc.
Effect of polymeric adsorbents on the production of sanguinarine by Papaver somniferum cell cultures
Authors: Williams Robert D. , Chauret Nathalie , Bédard Charles , Archambault Jean
Published: December 2004

The suitability of adsorbent polymeric resins, Amberlite XAD‐4 and XAD‐7 (Rohm and Hass, Inc.), was investigated for the accumulation of sanguinarine from Papaver somniferum cell cultures. The adsorption and desorption of sanguinarine from aqueous solution was most effective with XAD‐7. In addition to sanguinarine, the resins were found to absorb growth regulators and vitamins from the culture medium. Growth inhibition was overcome by delaying for ∼4 days resin addition after cell inoculation in fresh medium. Resin addition (5% wt/vol) to actively growing uneclicited cultures led to increases in sanguinarine production and release of 30% to 40% and 60%, respectively. The addition of resins to elicited cultures led to increases in alkaloid production of up to 50% to 85% with similar increases in alkaloid release as observed for nonelicited cells. Overall yield of sanguinarine increased from 21 mg · g biomass dry weight−1 (dw) for elicited cultures to more than 39 mg · gdw−1 when elicitation was combined with resin addition. Higher quantities of resin (10% to 20% wt/vol) increased marginally the release of sanguinarine into the medium, and on the resin, up to 85% of total production. The use of resin appears promesing for the development of a bioprocess for sanguinarine production by cultured plant cells. © 1992 John Wiley & Sons, Inc.
Tyrosinase reaction/chitosan adsorption for selectively removing phenols from aqueous mixtures
Authors: Payne Gregory F. , Sun Wei‐Qiang , Sohrabi Afshin
Published: December 2004

The goal of this work was to explore the technical feasibility of an enzymatic approach as an alternative to traditional approaches for phenol separations. Specifically, we examined a two‐step approach to selectively remove phenols from mixtures containing nonphenolic isomers. Our model solutes, of molecular formula C7H8O, were the phenol, cresol; the alkyl aryl ether, anisol; and the alcohol, benzyl alcohol. The first step is this two‐step approach employed the enzyme mushroom tyrosinase to selectively convert the phenolic, presumably to an o‐quinone product. The tyrosinase was specific for the phenol and was not observed to react with either the ether or the alcohol. The second step of this two‐step approach employed a sorbent of an appropriate surface chemistry to bind the products of the tyrosinase‐catalysed reaction of phenols. The sorbent used for this study was chitosan. Chitosan was observed to be unable to adsorb either nonphenol and was unable to adsorb unreacted cresol. However, Chittosan effectively adsorbs UV‐absorbing reaction products of the tyrosinase‐catalysed reaction of phenols. When mixtures of cresol and either anasol or benzyl alcohol were studied, the two‐step approach was effective for completely removing the phenolic without loss of either the ether or alcohol or the ether (i.e., phenols were removed with high separation factors).
Kinetic studies on urea hydrolysis by immobilized urease in a batch squeezer and flow reactor
Authors: Huang Ting‐Chia , Chen Dong‐Hwang
Published: December 2004

The immobilization of urease on the reticulated polyurethane foam, and the kinetic phenomenon of urea hydrolysis by the resulting immobilized urease in both batch squeezer and circulated flow reactors were studied. Urease was immobilized with bovine serum albumin and glutaraldehyde on polyurethane foam support of 7 to 15 μm thickness. The residual apparent activity of urease after immobilization was about 50%. The good hydrodynamic property and flexibility of polyurethane foam were retained in solution after immobilization. A modified biofilm reactor model was used to describe the kinetic phenomenon of urea hydrolysis in both batch squeezer and circulated flow reactors. The characteristic parameters of the reactor model for both bioreactors were obtained by combining the Rosenbrock optimization method, the Rungs–Kutta method, and the Newton–Raphson method. The best‐fit results were in good agreement with the experimental data. This study suggests another application of polyurethane foam in enzyme immobilization and immobilized enzyme reactors, which offers potential for practical applications in various bioreactors. © 1992 John Wiley & Sons, Inc.
An investigation of the diffusion‐limited growth of animal cells around single hollow fibers
Authors: Sardonini Charles A. , Dibiasio David
Published: December 2004

To compare modeling with experimental data of cell growth surrounding individual fibers, the growth profiles of hybridoma cells in the extracapillary space of single hollow fiber bioreactors were examined. Agarose was provided in the extracapillary space to provide support and minimize convection. By sacrificing bioreactors at various time intervals, the growth profiles of cells surrounding a single hollow fiber could be monitored with increasing time. Using photomicroscopy and viable staining, areas of viable and nonviable cell growth were examined at various stages of development ranging from initial seeding to stable growth conditions. Cells were found to act as nucleation sites for the growth of individual colonies within the agarose. Profiles at stable growth conditions resulted in a thick cell mass near the surface of the fiber wall followed by cell colonies of decreasing size with increasing radial distance. A simplified theoretical model for cell growth was developed using mass balance equations for substrate penetration into individual cell colonies as well as away from the wall of a single fiber. The resulting profiles derived from theory were compared with experiments and found to be in good agreement for entering oxygen concentrations of 5% and 20%. © 1992 John Wiley & Sons, Inc.
A unified stoichiometric model for oxidative and oxidoreductive growth of yeasts
Authors: Auberson Lillian C. M. , von Stockar U.
Published: December 2004

Yeasts degrade glucose through different metabolic pathways, where the choice of the pathway is dependent on the nature of the limitation in the various substrates. When oxygen is limiting in addition to glucose, yeasts often grow according to a mixture of oxidative and reductive metabolism. Oxygen may be limiting either by supply or by inherent biological restrictions such as the respiratory bottleneck in Saccharomyces cerevisiae or by both. A unified model incorporating both supply and biological limitations is proposed for the quantitative prediction of growth rates, consumption and production rates, as well as key metabolite concentrations during mixed oxidoreductive metabolism occuring as a result of such oxygen limitations. This simple unstructured model can be applied to different yeast strains while at the same time requiring a minimum number of measured parameters. “Estimators” are utilized in order to predict the presence of supply‐side or biological limitations. The values of these estimators also characterize the relative importance of oxidative to total metabolism. Results from the aerobic and oxygen‐limited chemostat cultures were used to corroborate the model predictions. During these experiments, the heat released by the yeast cultures was also monitored on‐line. The model correctly predicted the overall stoichiometry, steady‐state concentrations, and rates including heat dissipation rates measured in the various situations of oxygen limitations. Direct continuous measurements such as heat can be used in conjunction with the unified model for on‐line proces control. © 1992 John Wiley & Sons, Inc.