U. von Stockar

Characterization of an encapsulation device for the production of monodisperse alginate beads for cell immobilization

An encapsulation device, designed on the basis of the laminar jet break-up technique, is characterized for cell immobilization with different types of alginate. The principle of operation of the completely sterilizable encapsulator, together with techniques for the continuous production of beads from 250 microm to 1 mm in diameter, with a size distribution below 5%, at a flow rate of 1-15 mL/min, is described. A modification of the device, to incorporate an electrostatic potential between the alginate droplets and an internal electrode, results in enhanced monodispersity with no adverse effects on cell viability. The maximum cell loading capacity of the beads strongly depends on the nozzle diameter as well as the cells used. For the yeast Phaffia rhodozyma, it is possible to generate 700 microm alginate beads with an initial cell concentration of 1 x 10(8) cells/mL of alginate whereas only 1 x 10(6) cells/ml could be entrapped within 400 microm beads. The alginate beads have been characterized with respect to mechanical resistance and size distribution immediately after production and as a function of storage conditions. The beads remain stable in the presence of acetic acid, hydrochloric acid, water, basic water, and sodium ions. The latter stability applies when the ratio of sodium: calcium ions is less than 1/5. Complexing agents such as sodium citrate result in the rapid solubilization of the beads due to calcium removal. The presence of cells does not affect the mechanical resistance of the beads. Finally, the mechanical resistance of alginate beads can be doubled by treatment with 5-10 kDa chitosan, resulting in reduced leaching of cells.

The effect of yeast extract supplementation on the production of lactic acid from whey permeate by Lactobacillus helveticus.

SummaryBatch and continuous two-stage cultures have been conducted in order to determine the effect of yeast extract (YE) on the homolactic fermentation of whey permeate byLactobacillus helveticus. Supplementation with YE had a significant effet on lactic acid concentration, volumetric productivity, and substrate conversion, but not on lactic acid yield. Volumetric productivity in the first stage increased from 2 to 9 g l−1 per hour by increasing the YE concentration from 1.5 to 25 g l−1 At the same time conversion improved from 22% to 93% at a dilution rate of 0.2 h−1. The second stage demonstrated the effect of YE at a lower dilution rate (0.14 h−1. A high system conversion (97%) and a high final lactic acid concentration (40 g l−1) were achieved with 10 g l−1 YE.

Extractive bioconversion of 2-phenylethanol from L-phenylalanine by "Saccharomyces cerevisiae"

The bioconversion of l‐phenylalanine (l‐Phe) to 2‐phenylethanol (PEA) by the yeast Saccharomyces cerevisiae is limited by the toxicity of the product. PEA extraction by a separate organic phase in the fermenter is the ideal in situ product recovery (ISPR) technique to enhance productivity. Oleic acid was chosen as organic phase for two‐phase fed‐batch cultures, although it interfered to some extent with yeast viability. There was a synergistic inhibitory impact toward S. cerevisiae in the presence of PEA, and therefore a maximal PEA concentration in the aqueous phase of only 2.1 g/L was achieved, compared to 3.8 g/L for a normal fed‐batch culture. However, the overall PEA concentration in the fermenter was increased to 12.6 g/L, because the PEA concentration in the oleic phase attained a value of 24 g/L. Thus, an average volumetric PEA production rate of 0.26 g L−1 h−1 and a maximal volumetric PEA production rate of 0.47 g L−1 h−1 were achieved in the two‐phase fed‐batch culture. As ethanol inhibition had to be avoided, the production rates were limited by the intrinsic oxidative capacity of S. cerevisiae. In addition, the high viscosity of the two‐phase system lowered the kla, and therefore also the productivity. Thus, if a specific ISPR technique is planned, it consequently has to be remembered that the productivity of this bioconversion process is also quickly limited by the kla of the fermenter at high cell densities.

Does microbial life always feed on negative entropy? Thermodynamic analysis of microbial growth

Schrödinger stated in his landmark book, What is Life?, that life feeds on negative entropy. In this contribution, the validity of this statement is discussed through a careful thermodynamic analysis of microbial growth processes. In principle, both feeding on negative entropy, i.e. yielding products of higher entropy than the substrates, and generating heat can be used by microorganisms to rid themselves of internal entropy production resulting from maintenance and growth processes. Literature data are reviewed in order to compare these two mechanisms. It is shown that entropy-neutral, entropy-driven, and entropy-retarded growth exist. The analysis of some particularly interesting microorganisms shows that enthalpy-retarded microbial growth may also exist, which would signify a net uptake of heat during growth. However, the existence of endothermic life has never been demonstrated in a calorimeter. The internal entropy production in live cells also reflects itself in the Gibbs energy dissipation accompanying growth, which is related quantitatively to the biomass yield. An empirical correlation of the Gibbs energy dissipation in terms of the physico-chemical nature of the growth substrate has been proposed in the literature and can be used to predict the biomass yield approximately. The ratio of enthalpy change and Gibbs energy change can also be predicted since it is shown to be approximately equal to the same ratio of the relevant catabolic process alone.

Separation of binary mixtures by thermostatic sweeping gas membrane distillation: II. Experimental results with aqueous formic acid solutions

Aqueous solutions of formic acid have been experimentally investigated in a modified sweeping gas membrane distillation (SGMD) configuration. A thermostated sweeping gas was used in order to enhance the mass transfer performance. A new tubular module was designed and built for this purpose. The effects of the relevant process parameters on the permeate flux and selectivity have been studied. Experiments with pure water and pure formic acid were used to estimate certain parameters in the model. From these mass transfer coefficients, the fluxes and selectivity for aqueous formic acid mixtures have been calculated using the mathematical model previously described [J. Membr. Sci., 2001, in press]. The model predictions were compared with the experimental data and a good agreement between both flux values were obtained.

Optimal operation of fed-batch fermentations via adaptive control of overflow metabolite

The maximization of biomass productivity in the fed-batch fermentation of Saccharomyces cerevisiae is analyzed. Due to metabolic bottleneck, often attributed to limited oxygen capacity, ethanol is formed when the substrate concentration is above a critical value, which results in a decrease in biomass productivity. Thus, to maximize the production of biomass, the substrate concentration should be kept at the critical value. However, this value is unknown a priori and may change from experiment to experiment. A way to overcome this lack of knowledge is to allow the cells to produce a very small amount of ethanol. This way, the problem of maximizing the production of biomass is converted into that of regulating the concentration of ethanol, for which cell growth can be viewed as a perturbation. A novel adaptive control methodology based on the internal model principle is used to maintain the desired ethanol setpoint and reject the perturbation. Only a single parameter needs to be estimated on-line. Experimental results demonstrate the effectiveness of the proposed control methodology.

Inhibition aspects of the bioconversion of l-phenylalanine to 2-phenylethanol by Saccharomyces cerevisiae

The inhibitory impacts of the bioconversion of L-phenylalanine (L-Phe) to 2-phenylethanol (PEA), a very important natural aroma compd., on the metab. of Saccharomyces cerevisiae Giv 2009 were investigated in batch and chemostat cultures. The bioconversion was found to be completely growth assocd. and lead to a maximal final PEA concn. of 3.8 g/l of PEA. This was attained in a fed-batch procedure on glucose in order to prevent the formation of ethanol, which generally reduced the final achievable PEA concn. by its synergistic inhibitory effect. Chemostat cultures revealed that the bioconversion uncoupled the catabolism from anabolism of S. cerevisiae esp. under oxidative growth conditions and thereby reduced the crit. diln. rate Dcrit. In addn., higher specific oxygen uptake rates qO2 were found in the presence of the bioconversion at oxidative growth than the maximal respiratory capacity qO2max found in continuous cultures without bioconversion. [on SciFinder (R)]

Large-scale calorimetry and biotechnology

Heat dissipation in biotechnical processes is often a very conspicuous phenomenon in large scale bioreactors. These reactors could hence be considered as a form of calorimeter in that the heat dissipation rate could be measured without sophisticated probes or instrumentation. Such measurements are scarce in large scale fermenters, but literature exists on calorimetric measurements carried out in the laboratory under conditions closely resembling those maintained in industrial reactors. The calorimetric techniques used for such experiments are reviewed briefly, followed by a description of a few examples of the results obtained. Published reports are described which illustrate how the knowledge gained by such experiments could be applied to industrial bioprocesses in order to control the reactor, based on monitoring the heat dissipation rate of the culture.

Biological reaction calorimetry: development of high sensitivity bio-calorimeters

A review, with .apprx.43 refs. A review of different types of biol. reaction calorimetry systems currently used together with the operating principles is presented. The av. resoln. of these systems is approx. 20 to 1000 mW l-1, sufficient for studies of a wide range of cell culture processes. Poorly exothermic and endothermic processes require the development of even higher resoln. systems. To this end, the Mettler-Toledo RC1 calorimeter has been extensively studied to det. the factors which limit the resoln. By changing both the hardware and software, the resoln. has been increased to 2-5 mW l-1 for non-aerated processes and to 10-15 mW l-1 for aerated systems. The changes include a switchable elec. heater for the oil circulation thermostat, a new higher resoln. A/D board, PI controller and a thermostat reactor housing. The online measurement of the power input through agitation is proposed to be essential for low heat output biol. processes, even under conditions where the rheol. properties of the culture are not believed to be changing. The results show that it is possible to develop high-resoln. systems capable of operating under std. lab. bioreactor conditions; however, it is felt that the limits to the instrument resoln. have been attained and that the calorimetric signal resoln. is limited by the requirement of high agitation, nutrient feeds, gassing, pH control and other external effects which can only be overcome by heat-balancing methods. [on SciFinder (R)]

Separation of binary mixtures by thermostatic sweeping gas membrane distillation: I. Theory and simulations

A math. model describing a novel sweeping gas membrane distn. (SGMD) configuration is presented and the sepn. of two volatile components from binary mixts. by this membrane process is theor. investigated. SGMD configuration is modified by using a thermostated sweeping gas in order to enhance the driving forces decrease along the module. A Stefan-Maxwell-based model that includes vapor-liq. equil. and heat and mass transfer relations is used. This model that takes into account temp. and concn. polarization effects as well as temp. and concn. variation along the module length is employed to predict the flux and selectivity under the relevant operating conditions. Finally, a theor. evaluation of this particular configuration with respect to direct contact MD configuration is discussed. [on SciFinder (R)]