A model for growth of Saccharomyces cerevisiae containing a recombinant plasmid in selective media

A major problem in the use of plasmids as recombinant vectors is the problem of plasmid-free cell generation from plasmid shedding and subsequent growth. A common technique for controlling the population of plasmidfree cells is the use of selective media against these cells using an auxotrophic host and a plasmid that has the ability to produced the essential metabolite. A distributed model describing the growth of Saccharomyces cerevisiae containing a recombinant plasmid in selective media was developed. The model allows for growth and production of a metabolite by the plasmid-carrying strain and growth of the plasmid-free cells on resulting metabolite concentrations. Through a determination of system constants and numerical solution to the equations, experimental batch and continuous culture results for cell concentration transients could be simulated by the model. The results indicated that despite selective pressure, plasmid-free cell growth was significant.     

Improvement in separation characteristics of protein precipitates by acoustic conditioning

The effect of acoustic conditioning on the particle size distribution of isoelectric and calcium-ion-precipitated soya protein has been examined in low-residence-time chambers. In a previous study a beat frequency of 5 Hz obtained using a dual-source system of opposing vibrators was determined as giving optimal improvement in particle-settling characteristics for isoelectric soya protein precipitate. In this study the effect of amplitude of vibration, a measure of acoustic power input, and residence time of acoustic conditioning has been examined.Acoustic power input changed the flow pattern in the conditioning chamber from laminar streamline flow to a well-mixed, turbulent pattern. Such a mixing effect promoted the rapid aggregation of fine particles, a process that was modeled on the basis of orthokinetically controlled collisions. The rate of removal of fine particles due to acoustic conditioning was shown to be proportional to a mixing effect that was releated to the acoustic power dissipated per unit volume.The consequences of fine-particle aggregation on the centrifugal recovery of the precipitate are discussed.     

Mathematical analysis of two-phase mass transfer in a batch reactor for the chemical transformation of a steroid

A reactor is described for the conversion of the slightly water-soluble steroid testosterone (T) to 4-androstene-3, 17-dione (4-AD) by enzyme in the presence of excess cofactor. Since the enzyme is subject to substrate inhibition, reaction rates are strong functions of aqueous substrate concentration. High concentrations of the substrate, testosterone, per unit reactor volume are maintained within poly(dimethylsiloxane) beads that are suspended in the aqueous enzyme solution. Mass transfer (controlled by bead size, polymer to water volume ratio, enzyme loading) is used to control the degree and rate of conversion. The reactor dynamics are predicted over a wide range of reaction conditions. The product steroid is recovered in the polymeric beads from the enzyme solution.     

Enzymic interesterification of fats: Laboratory and pilot-scale studies with immobilized lipase from Rhizopus arrhizus

An immobilized lipase suitable for fat interesterification has been prepared by precipitation with acetone of a commercial lipase from Rhizopus arrhizus onto diatomaceous earth. As observed previously with a less active enzyme from Aspergillus sp., the interesterification activity was enhanced by addition of purified lipase or by high loadings of commercial enzyme. The interesterification activities reached maximum values in both cases. For immobilized preparations with purified enzyme, interesterification activity was also enhanced by the presence of a precoat of glutaraldehyde cross-linked commercial lipase. A 2.9-L column of immobilized lipase was used to interesterify batches of shea oleine (67 kg) and shea oil (40 kg). Little activity was lost processing shea oleine, but slow poisoning of the bed occurred when shea oil was fed to the column.     

Gibberellic acid by solid state fermentation: Consistent and improved yields

Five strains each of Gibberella fujikuroi and Fusarium monoliforme were screened to select G. fujikuroi P-3, a strain capable of giving consistent production of gibberellic acid (GA(3)) by solid state fermentation (SSF). The comparative production of GA(3) by SSF and submerged fermentation (SmF) indicated better productivity with the former technique. The accumulation of GA(3) was 1.626 times higher in the case of SSF. On the basis of available carbohydrates in the media, the percent conversions were 0.096 and 0.156 in SmF and SSF, respectively. The use of coarse wheat bran of the particle size of 0.3-0.4 cm resulted in an increase of 2.5 times in the yield of GA(3). The enrichment of commercial wheat bran with soluble starch gave enhanced accumulation to an extent of 3.5 times. The relation between GA(3) production and cell growth in SSF was similar to that encountered in SmF. The consistent and improved yields to a tune of 1.22 g GA(3) per kilogram dry moldy bran (DMB) establish the potential and feasibility of SSF for the production of GA(3) by G. fujikuroi P-3. On preliminary cost analysis, a net savings of about 60% and 50% on fermentation medium cost and the expenditure on down-stream processing, respectively, as compared to the presently employed SmF technique was evident.     

Conformational studies of A23187 with mono-, di- and trivalent metal ions by circular dichroism spectroscopy

CD studies carried out on A23187 indicate a solvent-dependent conformation for the free acid. Alkali metal ions were found to bind to the ionophore weakly. Divalent metal ions such as Mg2+, Ca2+, Sr2+, Ba2+ and Co2+ and trivalent lanthanide metal ions like La3+ were found to form predominantly 2:1 (ionophore-metal ion) complexes at low concentrations of metal ions, but both 2:1 and 1:1 complexes were formed with increasing salt concentration. Mg2+ and Co2+ exhibit similar CD behaviour that differs from that observed for the other divalent and lanthanide metal ions. The structure of 2:1 complexes involves two ligand molecules coordinated to the metal ion through the carboxylate oxygen, benzoxazole nitrogen and keto-pyrrole oxygen from each ligand molecule along with one or more solvent molecules. Values of the binding constant were determined for 2:1 complexes of the ionophore with divalent and lanthanide metal ions.