The influence of immobilization and reduced water activity on gaseous-alkene oxidation by Mycobacterium PY1 and Xanthobacter PY2 in a gas-solid bioreactor

Immobilization of Mycobacterium PY1 and Xanthobacter PY2 in alginate and in or on hydroculture has a minor influence on the maximum rate of oxidation of propene and ethane. The apparent K(m) values of the immobilized cells are slightly higher than those of the free cells, indicating the presence of diffusion limitation in the immobilized systems. Both bacterial strains rapidly lose their alkene-oxidizing activity when the water activity is decreased. This decrease in activity is so rapid that most of the activity is lost already when the pores of the pertinent supports are still filled with water. Therefore, it is not possible with this system to study the transition of mass transfer of substrates entirely in the water phase to mass transfer to substrates entirely through the gas phase.     

Immobilization of biocatalysts in thermogels using the resonance nozzle for rapid drop formation and an organic solvent for gelling

Summary The resonance nozzle immobilization technique was tested with thermo-hardening hydrogels (agar, gellan and -carrageenan) and compared with the conventional needle technique. After nozzling the warm gel solution, the formed droplets were caught in an ice-cold two-phase system consisting of an organic solvent (n-butylacetate, hexane or nonane) as top phase, in which gelling occurred, and aqueous medium as bottom phase. The cells used were yeast cells (Saccharomyces cerevisiae), bacterial cells (Mycobacterium aurum L1), plant cells (Tagetes minuta) and insect cells (Spodoptera frugiperda). The retention of the respiration activity was used as a criterion for the suitability of the conditions applied. The relatively polar solvent n-butylacetate with the low log P value gave the poorest activity retention, which is in agreement with experimentally validated theory. For all gels, the resonance nozzle technique proved to give satisfactory results, although the retention of respiration activity was generally lower than obtained with the needle technique.     

Stereospecific formation of 1,2-epoxypropane, 1,2-epoxybutane and 1-chloro-2,3-epoxypropane by alkene-utilizing bacteria

Resting cells of ethene grown Mycobacterium 2W produced 1,2-epoxypropane stereospecifically from propene as revealed by optical rotation, ¹H n.m.r. using a chiral shift reagent, and also by complexation gas chromatography involving a glass capillary column coated with an optically active metal chelate. The gas-liquid chromatography method allowed the rapid screening of 11 strains with regard to stereospecific formation of 1,2-epoxypropane, 1,2-epoxybutane and 1-chloro-2,3-epoxypropane. Bacteria grown on either ethene, propene or butadiene all predominantly produced the R form of 1,2-epoxypropane from propene and 1,2-epoxybutane from 1-butene while the strains tested for 1-chloro-2,3-epoxypropane production from 3-chloro-1-propene predominantly accumulated the S enantiomer.     

The magic-bead concept: an integrated approach to nitrogen removal with co-immobilized micro-organisms

This paper describes both qualitative and quantitative aspects of simultaneous autotrophic nitrification and heterotrophic denitrification by, respectively, the nitrifierNitrisomonas europaea and either of the denitrifiersPseudomonas denitrificans orParacoccus denitrificans co-immobilized in double-layer gel beads. The system is based on the establishment of well-defined oxic and anoxic zones within the cell supports and on physical separation of the nitrifying and denitrifying populations. Nitrification and denitrification rates were obtained from measured bulk concentrations and head-space analysis. The latter analyses showed that ammonia was primarily converted into molecular nitrogen. Nitrous oxide was not detected. High nitrogen removal rates (up to 5.1 mmol N m^–3 gel s^–1) were achieved in continuous reactors under aerobic conditions. The overall rate of nitrogen removal was controlled by the nitrifying step. The approach followed is, in principle, also suitable to the coupling of other oxidative and reductive bioprocesses having complementary metabolic routes. Two-stage bioconversion processes can be thus conducted as if single-staged, which results in more compact reactor systems.     

Silicone-stimulated anthraquinone production and release by Morinda citrifolia in a two-liquid-phase system

Summary A 2.5-fold increase in the release of intracellular anthraquinones was obtained by adding 1 ml L^-1 silicone A to suspension cultures of Morinda citrifolia. Cell growth and secondary metabolite production were not affected even at high silicone A concentrations. Performance of the silicone treatment in a two-liquid-phase system (5 ml n-hexadecane/50 ml medium) resulted in a 150% increase of the overall secondary metabolite productivity.     

Medium design based on stoichiometric analysis of microbial transglutaminase production by Streptoverticillium mobaraense

A stoichiometric model was developed for the application of medium design in microbial transglutaminase production by Streptoverticillium mobaraense. The model avoids dealing with all the metabolic reactions involved by simply lumping them into a single reaction. With the help of measurement results, an analysis of the nutrients' roles, and biochemical knowledge of the microorganism, all stoichiometric coefficients in the model were calculated. These coefficients were used for medium design. With this designed medium, microbial transglutaminase activity was increased fourfold, compared to that in the basal medium. (c) 1996 John Wiley & Sons, Inc.     

Hybridomas in a bioreactor cascade: modeling and determination of growth and death kinetics

Hybridomas were cultured under steady-state conditions in a series of two continuous stirred-tank reactors (CSTRs), using a serum-free medium. The substrate not completely converted in the first CSTR, was transported with the cells to the second one and very low growth rates, high death rates, and lysis of viable cells were observed in this second CSTR. These conditions are hardly accessible in a single vessel, because such experiments would be extremely time-consuming and unstable due to a low viability. In contrast to what is often observed in literature, kinetic parameters could thus be derived without the neccessity for extrapolation to lower growth rates. Good agreement with literature averages for other hybridomas was found. Furthermore, showing that the reactor series is a valuable research tool for kinetic studies under extreme conditions, the possibility to observe cell death under stable and defined steady-state conditions offers interesting opportunities to investigate apoptosis and necrosis. Additionally, a model was developed that describes hybridoma growth and monoclonal antibody production in the bioreactor cascade on the basis of glutamine metabolism. Good agreement between the model and the experiments was found.Abbreviation MAb Monoclonal antibody Nomenclature C _AConcentration of any (mol m^-3) component A D Dilution rate (s^-1) K _dDeath-rate constant (mol m^-3) K _lLysis-rate constant (mol m^-3) K _sMonod constant (mol m^-3) m Maintenance coefficient (mol cell^-1 s^-1) q Specific consumption (mol cell^-1 s^-1) or production rate t Time (s) X Cell concentration (cell m^-3) Y Yield coefficient (cell mol^-1) Greek symbols _d Specific death rate (s^-1) _l Specific lysis rate (s^-1) of viable cells _net Net specific growth (s^-1) rate _true True specific growth (s^-1) rate     

Quantitative Determination of the Spatial Distribution of Nitrosomonas europaea and Nitrobacter agilis Cells Immobilized in κ-Carrageenan Gel Beads by a Specific Fluorescent-Antibody Labelling Technique

A novel technique, combining labelling and stereological methods, for the determination of spatial distribution of two microorganisms in a biofilm is presented. Cells of Nitrosomonas europaea (ATCC 19718) and Nitrobacter agilis (ATCC 14123) were homogeneously distributed in a κ-carrageenan gel during immobilization and allowed to grow out to colonies. The gel beads were sliced in thin cross sections after fixation and embedding. A two-step labelling method resulted in green fluorescent colonies of either N. europaea or N. agilis in the respective cross sections. The positions and surface areas of the colonies of each species were determined, and from that a biomass volume distribution for N. europaea and N. agilis in κ-carrageenan gel beads was estimated. This technique will be useful for the validation of biofilm models, which predict such biomass distributions.     

Effects of diffusion limitation on immobilized nitrifying microorganisms at low temperatures

Activation energies of suspended and immobilized nitrifying bacteria were determined and compared to determine if diffusion limitation results in decreased sensitivity for temperature. The activation energy for the respiration activity of suspended Nitrosomonas europaea and Nitrobacter agilis was found to be 86.4 and 58.4 kJ mol(-1), respectively. The activation energy for oxygen diffusion in the support material, kappa-carrageenan, determined from the effect of temperature on the effective diffusion coefficient (D), was 17.2 kJ mol(-1). Consequently, the apparent actvation energy of diffusion limited cells should be lower. It was indeed shown that due to the effect of diffusion limitation and to temperature effects on the Monod constant K(s), the immobilized-cell activity was less sensitive to temperature. The apparent activation energy for immobilized Ns. europaea was between 28.6 and 94.2 kJ mol(-1) and for immobilized Nb. agilis between 1.4 and 72.9 kJ mol(-1), depending on the oxygen concentration and temperature. (c) 1995 John Wiley & Sons, Inc.