摘要:

AbstractThe effects of ethanol on reactor performance were studied in a small, 5‐cm packed height, “differential” type immobilized cell reactor. Lactose utilizing yeast cells,Kluyveromyces fragilis, were absorbed to a porous adsorbant sponge matrix in a gas continuous reactor. Step changes in the feed ethanol concentration to the column (10–130 g/L) were used to test the reactor response over extended periods of time (about 30–50 h per dosage level) followed by a return to basal zero inlet ethanol feed. Effluent cell density and effluent cell viability were measured at intervals. An inhibitory response in ethanol productivity to feed dosage ethanol levels above 20 g/L was detected almost immediately, with a near steady state response noted within 2.5 h of initiating the dosage. Feed ethanol levels above 50 g/L resulted in a subsequent gradual decrease in reactor productivity over time, which was associated with a decrease in the fraction of viable shed cells in the reactor effluent. The reactor response to a step removal of the ethanol inhibition was also monitored. Quick and complete rebounding of the fermentation rate to the original basal rate was noted following dosage concentrations of under 50 g/L ethanol. Recovery rates slowed following ethanol dosage levels above 50 g/L. Viable shed cell density improved overtime during the slow recovery periods. Growth rates (as determined by shed cell density) were more strongly inhibited than productivity. Growth responded more slowly to changes in ethanol environment as growth rates at 30 h fell to about 40% of the rates measured 7.5 h after initiation of a dosage level. It is concluded that ethanol contributions to cell injury and death (and consequent ICR performance degradation) may be more important than ethanol inhibition of productivity rates in the long‐term operation of immobilized cell reactors at ethanol concentrations

ISSN:0006-3592    

DOI:10.1002/bit.260361002

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1990

数据来源: WILEY

摘要:

AbstractThe viable fraction of immobilized cells in a bioreactor may be critical in predicting long‐term or steady‐state reactor performance. The assumption of near 100% viable cells in a bioreactor may not be valid for portions of immobilized cell reactors (ICRs) characterized by conditions resulting in appreciable death rates. A mathematical model of an adsorbed cell type ICR is presented in which a steady‐state viable cell fraction is predicted, based on the assumptions of no cell accumulation in the reactor and a random loss of cells from the reactor. Data on cell death rates, cell growth rates, and productivity rates as functions of temperature, substrate, and ethanol concentration for the lactose utilizing yeastK. fragilliswere incorporated into this model. The steady‐state reactor viable cell fraction as predicted by this model is a strong function of both temperature and ethanol concentration. For example, a stable 20% viable fraction of the immobilized cells is predicted in ICR locations experiencing continuous conditions of either 30 g/L ethanol at 40°C, or 95 g/L ethanol at 25°C. Steady‐state ICR “plug flow” concentration profiles and column productivities are predicted at three operating temperatures, 20, 30, and 40°C using two different models for ethanol inhibition of productivity. These profiles suggest that the reactor operating temperature should be low if higher outlet ethanol concentrations are desired. Three reactor design strategies are presented to maximize the viable cell fraction and improve long‐term ethanol productivity in ICR's: (1) reducing outlet ethanol concentrations, (2) rotating segments of an ICR between high and low ethanol environments, and (3) simultaneous removal of the ethanol produced from the react

ISSN:0006-3592    

DOI:10.1002/bit.260361003

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1990

数据来源: WILEY

摘要:

AbstractThe effect of reduced nutritional levels (particularly nitrogen source) for immobilizedK. fragilistype yeast were studied using a trickle flow, “differential” plug flow type reactor with cells immobilized by adsorption onto an absorbant packing matrix. Minimizing nutrient levels in a feed stream to an immobilized cell reactor (ICR) might have the benefits of reducing cell growth and clogging problems in the ICR, reducing feed preparation costs, as well as reducing effluent disposal costs. In this study step changes in test feed medium nutrient compositions were introduced to the ICR, followed by a return to a basal medium. Gas evolution rates were monitored and logged on a continuous basis, and effluent cell density was used as an indicator of cell growth rate of the immobilized cell mass. Startup of the reactor using a YEP medium showed a rapid buildup of cells in the reactor during the initial 110 h operation. The population density then stabilized at 1.6 × 1011cells/g sponge. A defined medium containing a complex mix of essential nutrients with an inorganic nitrogen source (ammonium sulfate) was able to maintain 90% of the productivity in the ICR as compared to the YEP medium, but proved unable to promote growth of the immobilized cell mass during startup. Experiments on reduced ammonium sulfate in the defined medium, and reduced yeast extract and peptone in YEP medium indicated that stable productivity could be maintained for extended periods (80 h) in the complete absence of any nutrients besides a few salts (potassium phosphate and magnesium sulfate). It was found that productivity rates dropped by 35–65% from maximal values as nitrogenous nutrients were eliminated from the test mediums, while growth rates (as determined by shed cell density from the reactor) dropped by 75–95%. Thus, nutritional deficiencies largely decoupled growth and productivity of the immobilized yeast which suggests productivity is both growth‐ and non‐growth‐associated for the immobilized cells. A yeast extract concentration of 0.375 g/L with or without 1 g/L ammonium sulfate was determined to be the minimum level which gave a sustained increase in productivity rates as compared to the nutritionally deficient salt medium. This represents a 94% reduction in complex nitrogenous nutrient levels compared to standard YEP batch medium (3 g/L YE and 3.

ISSN:0006-3592    

DOI:10.1002/bit.260361004

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1990

数据来源: WILEY

摘要:

AbstractAmino groups of trypsin (EC 3.4.21.4) were reductively alkylated in solid phase to obtain a surface‐active and biologically active enzyme in an o/w emulsion system. Trypsin adsorbed on a benzamidine–sepharose column was reductively alkylated withn‐octanal in the presence of sodium borohydride, i.e., trypsin‐C8. Activity of trypsin‐C8 againstNα‐benzoyl‐L‐arginine‐p‐nitroanilide was three times higher than that of native trypsin. Activities of trypsin and trypsin‐C8 against casein were almost the same. After incubating the trypsin solution at 40°C for 1 h, residual activities in the emulsion and solution systems were 64.2 and 57.4%, respectively. On the other hand, residual activities of native trypsin following incubation were 21.8% in the emulsion system and 33.2% in the solution system. Enhancement of trypsin‐C8 stability in the emulsion system may derive from interaction between the hydrophobic areas of trypsin‐C8 molecules and the hydrop

ISSN:0006-3592    

DOI:10.1002/bit.260361005

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1990

数据来源: WILEY

摘要:

AbstractThe growth and metabolism ofSaccharomyces cerevisiaewas studied in steady‐state chemostat cultures under conditions of scarce oxygen and excess glucose. The specific ethanol productivity and specific glucose uptake rate were stimulated by 50% within a narrow range of air/nitrogen mixtures to the fermentor. Fermentation was inhibited at slightly higher and lower air/nitrogen ratios, confirming similar results by previous investigators. This stimulation could not be caused by obvious mechanisms, such as the Pasteur or Crabtree effects. Since this maximum in the fermentation rate occurred in a steady‐state chemostat and at a constant dilution rate, the ATP yield of the culture necessarily attained a minimum. Thus, changes in the energetic efficiency of growth or the degree of wasting of ATP were surmised. The steady‐state biomass concentration at various oxygenation rates exhibited hysteresis phenomena. Ignition and extinction of the biomass concentration occurred as critical oxygen feed rates were passed. The hysteresis was prevented by adding yeast extract to or reducing the antifoam concentration in the medium. These medium alterations had the simultaneous effect of stimulating the fermentation rate, suggesting that ATP has a critical role in dictating the biomass concentration in micro‐aerobic culture. Silicone polymer antifoam was found to stimulate glycerol production at the expense of ethanol production, having consequences for the energy generation and the biomass concentration of the

ISSN:0006-3592    

DOI:10.1002/bit.260361006

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1990

数据来源: WILEY

摘要:

AbstractIntracellular metabolite concentration and enzyme activity measurements were made to explain the new metabolic and growth phenomena seen in the micro‐aerobic, continuous yeast cultures described in Part I. The results of these assays suggested mechanisms for the observed maximum in the specific ethanol productivity as a function of the oxygen feed rate, changing ATP yields, the effects of antifoam, and the sharp changes in the biomass concentration with small changes in the oxygenation. Measured were the intracellular concentrations of ATP, NADH, glucose 6‐phosphate, pyruvate, glycerol, and ethanol, and the activities of hexokinase and alcohol dehydrogenase. Rate‐limiting steps were identified by the accumulation of metabolites upstream and the depletion of metabolites downstream of the step.A potential mechanism for the stimulation of fermentation with decreasing oxygenation was an activation of glucose transport by an accumulating intracellular ATP concentration. The inhibition of fermentation at yet lower oxygenation rates may have been caused by the continued accumulation of ATP to the point that the glycolytic kineses were inhibited. A mechanism for the changing ATP yields and intracellular ATP concentration proposed the existence of ATPases or ATP waste reactions stimulated by both oxygen and ATP. Antifoam had the effect of decreasing the resistance for glycerol transport out of the cell. The resulting stimulation of glycerol production and inhibition of ethanol production decreased the intracellular ATP content. Finally, intracellular ethanol was found not to accumulate to levels of higher than the extracellular concentr

ISSN:0006-3592    

DOI:10.1002/bit.260361007

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1990

数据来源: WILEY

摘要:

AbstractMathematical models of the catabolic pathways, the utilization and waste of ATP, and the factors affecting yeast growth in a micro‐aerobic chemostat are presented. The models incorporate the intracellular metabolite and enzyme activity assays performed in Part II to explain the unusual macroscopic chemostat behaviors reported in Part I. The catabolic model successfully predicts a maximum in the specific ethanol productivity as a function of the intracellular ATP concentration. The ATP balance model enables the prediction of the intracellular ATP concentration and the ATP yield for given dissolved oxygen concentrations. Finally, in the context of a growth model, singularity theory provides a framework to explain the transition observed in Part I between hysteresis and the monotonic biomass versus oxygenation profiles in response to changes in the nutrient composition. The models serve to organize data and to concretely express proposed metabolic mechanisms and cause–effect hypotheses. The model is only applicable to the micro‐aerobic and excess glucose conditions encountered in this

ISSN:0006-3592    

DOI:10.1002/bit.260361008

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1990

数据来源: WILEY

摘要:

AbstractThis article presents an introduction to the use of neural network computational algorithms for the dynamic modeling of bioprocesses. The dynamic neural model is used for the prediction of key fermentation variables. This relatively hew method is compared with a more traditional prediction technique to judge its performance for prediction. Illustrative simulation results of a continuous stirred tank fermentor are used for this comparison. It is shown that neural network models are accurate with a certain degree of noise immunity. They offer the distinctive ability over more traditional methods to learn very naturally complex relationships without requiring the knowledge of the model structure.

ISSN:0006-3592    

DOI:10.1002/bit.260361009

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1990

数据来源: WILEY

摘要:

AbstractHybridoma cells (S3H5/γ2bA2) were cultivated in spinner flasks with 1% serum media and serum‐free media. Monoclonal antibody productivity was maintained in 1% serum media. However, cells in serum‐free media showed a decrease in antibody productivity, and it completely disappeared in IMDM‐based low protein medium. This loss of antibody productivity was not observed when the cells were immobilized in alginate beads. In fact, immobilization enhanced the specific MAb produc

ISSN:0006-3592    

DOI:10.1002/bit.260361010

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1990

数据来源: WILEY

摘要:

AbstractThe production of IGF‐1 (insulin‐like growth factor 1), expressed inEscherichia colias a secreted fusion protein with affinity for the Fc region of IgG, was monitored automatically during fermentations. A sampling device was used to automatically inject filtered culture medium from the fermentor onto a small affinity column (IgG Sepharose® Fast Flow) connected to a chromatographic system. The area of the eluted peak was proportional to the concentration of the fusion protein. The relationship was linear over the range 25–630 μg/mL with relative standard deviation of around 1% at the higher concentrations. Samples could be monitored automatically every half hour during fermentation (48 h). The method of analysis is nondestructive, allowing further analysis of product quality. A complete evaluation of the monitoring system is described. With this system, fermentations based on the described expression system can be optimized on the basis of product concentration; this will lead to more effective fermentations and higher product yields. It should also be possible to monitor other secreted products with this system by using other affini

ISSN:0006-3592    

DOI:10.1002/bit.260361011

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1990

数据来源: WILEY