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Biomedical subjects

C L Cooney

Publications and source records attributed to C L Cooney.

17 recordsLinked to original sources
PubMed1992

Fermentation monitoring.

Fermentation monitoring continues to be the focus of much research. Over the last year, important strides were made in improving bioprocess monitoring using NADH fluorescence, viscosity, affinity techniques, enzyme and microbial sensors, calorimetry, flow injection analysis and bioluminescence. Better fermentation monitoring is important for improving understanding, operation, development and control of the process. We expect progress in these areas of research to continue. In addition, we highlight some non-conventional approaches.

Biosensing Techniques↗
PubMed1990

Optimization and simulation of continuous affinity-recycle extraction (care).

Simulation and optimization of continuous affinity recycle extraction (CARE), a protein purification unit operation based on protein adsorption to solid phase adsorbents, is described in this paper. Rather than packing conventional adsorbent particles in a fixed bed (column), solid/liquid contact is carried out in well-mixed reactors. Continuous operation is achieved by recirculation of the adsorbent particles between two or more contactors. The feasibility of this purification scheme was established with the recovery and isolation of the enzyme beta-galactosidase from E.coli, using the affinity support PABTG/Agarose. A mathematical model describing system performance was developed. The mathematical model was used to optimize several facets of the system design and operation. The base two-stage contractor design was modified by the addition of an intermediate wash stage as well as the incorporation of multiple adsorption stages. These design modifications serve to increase purification, concentration and recovery while utilizing the same amount of adsorbent. The methodology for defining and optimizing objective functions was developed and experimentally validated. Finally, optimum system start-up protocols, minimizing the time required to reach steady-state operation, were developed and experimentally validated. The impact of early introduction of adsorptive purification in a downstream processing sequence, with CARE, was evaluated and is described. Through the early introduction of a highly specific adsorptive step, significant purification is achieved simultaneously with clarification and concentration. In addition, purification performance in CARE was contrasted with that achievable in conventional column chromatography.

Chromatography, Affinity↗
PubMed1978

Acetate kinase production by Escherichia coli during steady-state and transient growth in continuous culture.

The synthesis of acetate kinase by Escherichia coli ATCC 9637 was studied during growth in anaerobic continuous cultures under steady-state and transient conditions. During growth in anaerobic, glucose-limited chemostats, acetate kinase synthesis was linearly associated with growth. Two types of non-steady-state transients were studied: the perturbation in one was the addition of glucose alone, and, in the second, glucose plus Casamino Acids. During the nutritional shift-up in the second case, but not in the first, the instantaneous specific acetate kinase activities and specific synthesis rates exceeded pre- and postshift values. Trajectory curves demonstrated that the increase in specific activity remained within the bounds of values obtainable under steady-state conditions with minimal and Casamino Acids media. Specific synthesis rates, however, greatly exceeded steady-state values. Enzyme yield values on glucose after the transient nutritional shift-up increased up to fivefold. Active protein synthesis is shown to be necessary to achieve the enhanced specific synthesis rates and enzyme yields. The results from these transient responses are discussed in terms of a conceptful model for metabolic regulation.

Acetates↗
PubMed1977

Computer-aided material balancing for prediction of fermentation parameters.

Despite the importance of biomass as a parameter in fermentation processes, there are no commercially available sensors suitable for its measurement. An indirect approach for the assessment of biomass concentration can be based on material balances and on the direct monitoring of fermentation parameters for which there are established sensors (e.g., gaseous oxygen and carbon dioxide). As a consequence, this method requires no assumption of cellular yield coefficients or rate constants. This approach is also readily adaptable to general use since it requires only some knowledge of the compositions of the substrate, cells, and noncellular products.

Carbon Dioxide↗
PubMed1977

Computer-aided baker's yeast fermentations.

The economics of yeast production depend heavily upon the cellular yield coefficient on the carbon source and the volumetric productivity of the process. The application of an on-line computer to maximize these two terms during the fermentation requires a continuous method of measuring cell density and growth rate. Unfortunately, a direct sensor for biomass concentration suitable for use in industrial fermentations is not available. Material balancing, with the aid of on-line computer monitoring, offers an indirect method of measurement. Laboratory results from baker's yeast production in a 14-liter fermentor (with a PDP-11/10 computer for on-line analyses) show this indirect measurement technique to be a viable alternative. From the oxygen uptake and carbon dioxide production data, gas flow rate, and ammonia addition rate, the cell density during the fermentation has been estimated and found to compare well with actual fermentation data.

Ammonia↗
PubMed1977

Theoretical conversion yields for penicillin synthesis.

The efficiency of conversion of the carbon-energy source to product is of primary importance in many fermentation processes. In order to assess the efficiency of a process, one must know how close the actual conversion yield is to the theoretical maximum. Theoretical conversion yields are useful, therefore, as guides in improving a process. This knowledge is particularly important today because the cost of raw materials is rapidly rising. In this study, the biochemical pathway of penicillin synthesis was used to estimate the theoretical yield of penicillin from glucose, ammonia, and sulfate. These values are compared with experimental data from the literature. An analysis of the role of glucose in the synthesis of cell mass and penicillin and in the maintenance of cells makes it possible to assess the efficiency of carbon-source utilization and to direct further advances in penicillin fermentations.

Ammonia↗
PubMed1977

Distribution of dextransucrase in Streptococcus mutans and observations on the effect of soluble dextran on dextransucrase activities.

Total and insoluble dextransucrase activities were measured in cell-associated and supernatant fractions of Streptococcus mutans GS-5 grown in several media. Although the amount of cell-associated and supernatant activity varied greatly as a function of medium, the total activity appeared constant. The distribution of dextransucrase could be altered without changing the total dextransucrase activity. This indicates that the distribution of the enzyme can be regulated independently of its synthesis. Strain GS-5 had significant cell-associated activity in media devoid of sucrose. In all cases, the ratio of insoluble to total dextransucrase activity was higher in the cell-associated fractions than in the cell-free supernatants. It is also demonstrated that exogenous soluble dextran caused a decrease in insoluble dextransucrase activity and an increase in soluble dextransucrase activity in both the cell-associated fraction and the culture supernatant. The stimulation of soluble dextran-synthesizing activity was not due to de novo synthesis. The inhibition of insoluble dextran-synthesizing activity is shown to be noncompetitive. These results support a physical rather than metabolic mechanism for the effect of soluble dextran on dextransucrase activities.

Chloramphenicol↗
PubMed1977

Measurement and synthesis of insoluble and soluble dextran by Streptococcus mutans.

Total and insoluble dextransucrase activities of 10 strains of oral streptococci were measured by a modified filter disk assay. Strains that were nonadherent to hard surfaces had only low levels of insoluble dextransucrase activity. A physical rather than metabolic mechanism is suggested to explain the decreased insoluble and increased soluble activities observed when dextran T-10 is added to the media.

Bacteriological Techniques↗
PubMed1976

Experiments and calculations concerning a thermal enzyme probe.

A simple device capable of measuring almost any reactant in an enzyme-catalyzed reaction is created when an enzyme is immobilized onto one thermal sensor of a differential thermometer. Experiments are described in which two thermistors, one bare and one coated with immobilized enzyme, are immersed in a well-stirred solution. The response of this device to increases in glucose-ATP concentration was observed using hexokinase (ATP:D-hexose 6-phosphotransferase, EC 2.7.1.1), and to increases in glucose concentration using glucose oxidase (beta-D-glucose:oxygen 1-oxidoreductase, EC 1.1.3.4). A simple model is presented whose predictions are in reasonable agreement with the experimental results.

Enzymes, Immobilized↗
PubMed1976

Transient response of Enterobacter aerogenes under a dual nutrient limitation in a chemostat.

Utilizing a chemostat with a dual nutrient limitation of nitrogen and phosphate, we examined the transient response of the culture following a pulse of one of the limiting nutrients (ammonia). This method provided quantitative evidence that cells can be grown under dual nutrient limitation. Furthermore, the pattern of response was consistent with the hypothesis that phosphate limitation restricts nucleic acid synthesis in the cell and that nitrogen limitation restricts protein synthesis. The net result is that under a phosphate limitation there is a restricted biosynthetic capacity which we feel is closely associated with the RNA content of the cell.

Culture Media↗
PubMed1976

Application of dynamic calorimetry for monitoring fermentation processes.

The rate of heat evolution (kcal/liter-hr) in mycelial fermentations for novobiocin and cellulase production with media containing noncellular solids was measured by an in situ dynamic calorimetric procedure. Thermal data so obtained have proved significant both in monitoring cell concentration during the trophophase (growth phase) and in serving as a physiological variable in the fermentation process. The validity of this technique has been demonstrated by closing the overall material and energy balances. The maintenance energy in a batch fermentation can be calculated by integrating heat evolution data. This integration method is applicable to a fermentation lacking a precise cell growth curve. The maintenance coefficient, obtained for the novobiocin fermentation by Streptomyces niveus, is equal to 0.028 g glucose equivalent/g cell-hr. The production of novobiocin in the idiophase (production phase) also correlates well with the amount of energy catabolized for maintenance and this results in an observed conversion yield of glucose to novobiocin of 11.8 mg of novobiocin produced per gram of glucose catabolized. A new physiological variable, kilocalories of heat evolved per millimole of oxygen consumed, has been proposed to monitor the state of cells during the fermentation. This method may provide a simple way to monitor on-line shifts in the efficiency of cell respiration and changes in growth yields during a microbial process.

Calorimetry↗
PubMed1976

Production of gramicidin S synthetases by Bacillus brevis in continuous culture.

The effects of different nutrient limitations on the production of the two enzymes of gramicidin S biosynthesis were studied during continuous culture of Bacillus brevis. Gramicidin S synthetases I and II were produced in the chemostat under carbon, nitrogen, phosphorus or sulphur limitation. The growth rate, rather than the nature of the limitation, was the major controlling factor in regulating the level of the gramicidin S synthetases. Synthetase production was low at high dilution rates (0.45 to 0.50 h-1) but increased as the dilution rate was lowered. The highest specific activities occurred at dilution rates that were different for each type of limitation: 0.40 h-1 for nitrogen, 0.32 h-1 for carbon, 0.24 h-1 for sulphur and 0.20 h-1 for phosphorus. Phosphorus limitation gave the highest specific activities. At low dilution rates (0.10 to 0.15 h-1), enzyme activities were again low. Sporulation occurred under carbon limitation, but at a lower dilution rate than that which supported optimal gramicidin S synthetase formation. The specific productivity of the synthetases in the chemostat was higher than the highest productivity obtained in batch growth.

Amino Acid Isomerases↗
PubMed1976

Growth of Enterobacter aerogenes in a chemostat with double nutrient limitations.

The behavior of Enterobacter aerogenes during growth in chemostats limited by single and double nutrient restrictions was examined. On the assumption that different essential nutrients act to limit growth in different ways, we selected pairs of nutrients likely to affect different aspects of metabolism. Results show that macromolecular cell composition can be controlled by using more than one nutrient restriction. The polysaccharide content of the cells is readily manipulated by the ratio of carbon to nitrogen in the inlet nutrients. Also, at low dilution rates, ratios of protein to ribonucleic acid are dependent on the ratio of phosphate to nitrogen in the input nutrients. An examination of both acetic acid and metabolite production (as measured by ultraviolet absorbance of culture filtrates) showed that accumulation of these products was dependent on both dilution rate and type of nutrient limitation(s). These results were examined in terms of the problems of translation of batch to continuous culture processes and the use of selected nutrient limitations to control noncellular product formation.

Ammonia↗
PubMed1975

Thermophilic anaerobic digestion of solid waste for fuel gas production.

Anaerobic digestion offers a potential means of converting organic solid waste into fuel gas and thereby provide a supplemental and readily utilizable source of energy. We are particularly interested in the use of thermophilic digestion over a mesophilic operation for it can achieve higher rates of digestion, greater conversion of waste organics to gas, faster solid-liquid separation, and minimization of bacterial and viral pathogen accumulation. Our results comparing mesophilic (37 degree C) and thermophilic (65 degree C) anaerobic digestion of domestic solid waste confirm the increased rate and conversion of waste to methane. In addition, utilizing radioactive labeling of glucose and acetic acid, we have measured the volumetric rates of volatile acid production and disappearance under both mesophilic and thermophilic conditions.

Anaerobiosis↗