Biomedical subjects

J M Scharer

Publications and source records attributed to J M Scharer.

9 recordsLinked to original sources

PubMed2005

Medium optimization for hen egg white lysozyme production by recombinant Aspergillus niger using statistical methods.

Statistics-based experimental design was used to investigate the effect of medium components (starch, peptone, ammonium sulfate, yeast extract, and CaCl2.2H2O) on hen's egg white lysozyme production by Aspergillus niger HEWL WT-13-16. A 2(5-1) fractional factorial design augmented with center points revealed that peptone, starch, and ammonium sulfate were the most significant factors, whereas the other factors were not important within the levels tested. The method of steepest ascent was used to approach the proximity of optimum. This task was followed by a central composite design to develop a response surface for medium optimization. The optimum medium composition for lysozyme production was found to be: starch 34 g L-1, peptone 34 g L-1, ammonium sulfate 11.9 g L-1, yeast extract 0.5 g L-1, and CaCl2.2H2O 0.5 g L-1. This medium was projected to produce, theoretically, 212 mg L-1 lysozyme. Using this medium, an experimental maximum lysozyme concentration of 209+/-18 mg L-1 verified the applied methodology.

Algorithms↗

PubMed2004

Fed-batch optimization of alpha-amylase and protease-producing Bacillus subtilis using Markov chain methods.

A stoichiometry-based model for the fed-batch culture of the recombinant bacterium Bacillus subtilis ATCC 6051a, producing extracellular alpha-amylase as a desirable product and proteases as undesirable products, was developed and verified. The model was then used for optimizing the feeding schedule in fed-batch culture. To handle higher-order model equations (14 state variables), an optimization methodology for the dual-enzyme system is proposed by integrating Pontryagin's optimum principle with fermentation measurements. Markov chain Monte Carlo (MCMC) procedures were appropriate for model parameter and decision variable estimation by using a priori parameter distributions reflecting the experimental results. Using a simplified Metropolis-Hastings algorithm, the specific productivity of alpha-amylase was maximized and the optimum path was confirmed by experimentation. The optimization process predicted a further 14% improvement of alpha-amylase productivity that could not be realized because of the onset of sporulation. Among the decision variables, the switching time from batch to fed-batch operation (t(s)) was the most sensitive decision variable.

Algorithms↗

PubMed2002

Bioprocessing strategies for improving hen egg-white lysozyme (HEWL) production by recombinant Aspergillus niger HEWL WT-13-16.

Hen egg-white lysozyme (HEWL) production by recombinant Aspergillus niger HEWL WT-13-16 from a cDNA under the control of the A. niger glucoamylase promoter was used as a model system. The fungal mycelium was either immobilized on porous Celite 560 micro-carrier or grown in suspension as pelleted and dispersed forms. The objective was to reduce the protease activity that adversely affects the expressed HEWL. Free suspension culture at uncontrolled pH served as the benchmark. The control of pH during growth at pH 4.0 gave rise to a greater than five-fold reduction of protease activity in suspension culture. An additional 38.5% decrease in protease activity was achieved in mycelial-pellet cultures in comparison to a 40.9% decrease in protease activity obtained with Celite 560 beads in an airlift vessel at controlled pH. The specific HEWL yields were 5.8, 5.0 and 4.1 mg/g dry wt. for the free suspension, mycelial-pellet, and Celite-560-immobilized cultures, respectively.

Animals↗

PubMed2001

Peptidases affecting recombinant protein production by Streptomyces lividans.

The influence of peptidases on human interleukin-3 (rhIL-3) production by a recombinant Streptomyces lividans strain was investigated. The bacterium produced several general peptidases and tripeptidyl peptidases compromising the authenticity of rhIL-3. The level of peptidases depended on growth morphology. Growing S. lividans as compact pellets successfully reduced peptidase activity. Maximum general peptidase activity in pellet culture was delayed after maximum rhIL-3 concentration was achieved. The activity of the tripeptidyl peptidase was product (rhIL-3) associated.

Aminopeptidases↗

PubMed1997

Plasmid instability kinetics in continuous culture of a recombinant Saccharomyces cerevisiae in airlift bioreactor.

Plasmid instability of a recombinant Saccharomyces cerevisiae C468/pGAC9 (ATCC 20690) was examined during continuous culture in a nonselective medium in an airlift bioreactor. The recombinant strain contained a 2-micron based shuttle vector pGAC9 and expresses Aspergillus awatnori glucoamylase gene under the control of the yeast enolase I (ENO1) promoter. The changes in the fraction of plasmid-bearing cells and glucoamylase activity followed first-order kinetics. Expressed as a function of time, the decay rates of both the plasmid-bearing cell fraction and glucoamylase expression increased with increasing dilution rates. If expressed as a function of cell generations, the decay rates were nearly constant over the dilution rates tested. The results indicated that the growth rate difference between plasmid-bearing and plasmid-free cells was negligible. This was probably due to the low copy number of the 2-micron based yeast shuttle vector. Thus the contribution of preferential growth to apparent plasmid instability was negligible. A novel numerical method is proposed to evaluate the parameters related to plasmid stability. The estimated values of probability of plasmid loss (P = 0.0499) were nearly constant at different dilution rates. No significant effect of growth rates on plasmid instability was observed. The proposed kinetics agreed well with experimental observations.

Bioreactors↗

PubMed1992

Design and performance of a trickle-bed bioreactor with immobilized hybridoma cells.

A trickle-bed system employing inert matrices of vermiculite or polyurethane foam packed in the downcomer section of a split-flow air-lift reactor has been developed for hybridoma culture to enhance antibody productivity. This quiescent condition favoured occlusion and allowed the cells to achieve densities twelve fold greater (12.8 x 10(6) cells/ml reactor for polyurethane foam) than in free cell suspension. The reactor was operated in a cyclic batch mode whereby defined volumes of medium were periodically withdrawn and replaced with equal volumes of fresh medium. The pH of the medium was used as the indicator of the feeding schedule. Glucose, lactate and ammonia concentrations reached a stationary value after 5 days. With vermiculite packing, a monoclonal antibody (MAb) concentration of 2.4 mg/l was achieved after 12 days. The MAb concentration declined then increased to a value of 1.8 mg/l. In the polyurethane foam average monoclonal antibody (MAb) concentrations reached a stationary value of 1.1 mg/l in the first 20 days and increased to a new stationary state value of 2.1 mg/l for the remainder of the production. MAb productivity in the trickle-bed reactor was 0.3 mg/l.d (polyurethane foam) and 0.18 mg/l.d (vermiculite) in comparison to 0.12 mg/l.d for free cell suspension. This trickle-bed system seems to be an attractive way of increasing MAb productivity in culture.

Alanine↗

PubMed1988

Media for hybridoma growth and monoclonal antibody production.

For the economical production of monoclonal antibodies (MAbs), the cell-culture medium must be optimized for three different phases: growth of the hybridomas, MAb productivity of the hybridomas, and MAb purification or downstream processing. Medium improvements are necessary to meet these requirements for large-scale MAb production. Information bearing on this issue is being addressed in two research areas, cell biology and biochemical engineering, and is reviewed in this article.

Journal Article↗

PubMed1987

Microbial desulphurization of heavy oils and bitumen.

Most oil producing countries have extensive reserves of heavy oil and bitumen. As easily accessible sources of conventional crudes decline, these reserves will become more important in supplementing the energy requirements. Heavy oil and bitumen are highly viscous and contain 3 to 6% sulphur. These objectionable quantities of sulphur must be removed before being acceptable as refinery feedstock. This paper addresses the potential of biological desulphurization of heavy oil and bitumen. The aerobic and anaerobic processes to remove organic as well as inorganic sulphur have been reviewed. To date, most studies were performed with model substrates, particularly dibenzothiophene (DBT) in a synthetic medium. Early work concerned with the isolation of microorganisms, identification and characterization of intermediate metabolites, and the development of growth media. No commercially viable process has emerged since the engineering details of the process have not been addressed conclusively. Due to high utility and catalyst cost conventional hydrodesulphurization processes are reported to be uneconomic in case of high sulphur oils. Microbial desulphurization, on the other hand, appears to be promising due to the inherent low energy requirement. This process may become more attractive by the application of genetically modified bacteria and improvements in bioreactor design.

Journal Article↗

PubMed1968

Growth and metabolism of live vaccine strain of Pasteurella tularensis.

The growth and metabolism of the live vaccine strain of Pasteurella tularensis in different media were investigated. Maximal growth was observed in a medium containing a sulfuric acid digest of casein as amino acid source. Amino acid metabolism produced considerable ammonia, and the rate of ammonia evolution was directly proportional to the growth rate. The most likely route for amino acid breakdown is nonspecific oxidative deamination.

Amino Acids↗