Staff Publications

Staff Publications

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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

    Full text documents are added when available. The database is updated daily and currently holds about 240,000 items, of which 72,000 in open access.

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    Fermentation characteristics of yeasts isolated from traditionally fermented masau (Ziziphus mauritiana) fruits
    Nyanga, L.K. ; Nout, M.J.R. ; Smid, E.J. ; Boekhout, C. ; Zwietering, M.H. - \ 2013
    International Journal of Food Microbiology 166 (2013)3. - ISSN 0168-1605 - p. 426 - 432.
    saccharomyces-cerevisiae - volatile compounds - wine yeasts - immobilized cells - grape varieties - zimbabwe - strains - metabolism - acid - evolution
    Yeast strains were characterized to select potential starter cultures for the production of masau fermented beverages. The yeast species originally isolated from Ziziphus mauritiana (masau) fruits and their traditionally fermented fruit pulp in Zimbabwe were examined for their ability to ferment glucose and fructose using standard broth under aerated and non-aerated conditions. Most Saccharomyces cerevisiae strains were superior to other species in ethanol production. The best ethanol producing S. cerevisiae strains, and strains of the species Pichia kudriavzevii, Pichia fabianii and Saccharomycopsis fibuligera were tested for production of flavor compounds during fermentation of masau fruit juice. Significant differences in the production of ethanol and other volatile compounds during fermentation of masau juice were observed among and within the four tested species. Alcohols and esters were the major volatiles detected in the fermented juice. Trace amounts of organic acids and carbonyl compounds were detected. Ethyl hexanoate and ethyl octanoate were produced in highest amounts as compared to the other volatile compounds. S. cerevisiae strains produced higher amounts of ethanol and flavor compounds as compared to the other species, especially fatty acid ethyl esters that provide the major aroma impact of freshly fermented wines. The developed library of characteristics can help in the design of mixtures of strains to obtain a specific melange of product functionalities. Keywords: Masau juice; Fermentation; Yeast; Volatile compounds; Flavor; Wine
    Nitrification by artificially immobilized cells : model and practical system
    Leenen, E.J.T.M. - \ 1997
    Agricultural University. Promotor(en): J. Tramper; R.H. Wijffels. - S.l. : Leenen - ISBN 9789054856740 - 168
    nitrificatie - cellen - immobilisatie - geïmmobiliseerde cellen - nitrification - cells - immobilization - immobilized cells

    The research described in this dissertation started after extended studies on the fundamental aspects of the growth of immobilized nitrifying cells. In these previous studies dynamic models were developed and experimentally validated, which resulted in knowledge about which steps determine the rate of the nitrification process and to which parameters the immobilized-cell process is sensitive. Furthermore, it showed that this system may be interesting as a practical system as well.

    The aim of the work described in this thesis was to obtain more insight in the complex interactions in the nitrification process with artificially immobilized cells in air-lift loop reactors. The knowledge obtained may form the basis for a rational design of compact systems for practical applications, e.g. nitrification of domestic wastewater. This knowledge was built up by developing dynamic models, by studying the applicability and mechanical stability of the immobilization support and by doing experiments in domestic wastewater.

    In Chapter 1 an overview of the previous studies and the approach, followed in our group for the development of a process with immobilized cells, are given. The subjects in this Chapter provide the starting points of the following chapters, which are arranged in three themes: dynamics, supports and application.

    The first theme, the dynamics of immobilized nitrifying cells entrapped in carrageenan, is described in Chapter 2 and 3. The results in Chapter 2 clearly show the reduced temperature sensitivity of immobilized nitrifying cells caused by diffusion limitation. In Chapter 3 is described that, for example at low substrate concentrations and low hydraulic retention times, the death rate of cells is an important parameter. If cells die as a result of e.g. a low substrate loading, the process reacts slower to changes in substrate concentrations than if all cells remain viable.

    The second theme (Chapter 4 and 5) deals with the selection and characterization of support materials for cell immobilization. In order to establish which support materials are the most suitable, characteristics of several natural and synthetic materials have been determined and compared, Natural gel materials, like alginate and carrageenan, have a mild immobilization procedure such that few cells die and they grow well in these supports. These supports, however, appeare to be soluble, biodegradable and sensitive to abrasion.

    Synthetic gels, on the contrary, have better mechanical properties, but the immobilization conditions are less mild resulting in low biomass retention. For application of entrapped nitrifying cells synthetic gels, like polyethylene glycol, polyvinyl alcohol and polycarbamoyl sulphonate, are, however, promising (Chapter 4). In Chapter 5 the relevance of rheological properties of gel beads for their mechanical stability is described. It is concluded that tests based on rupture of 'virgin' beads does not provide relevant information on the mechanical stability and sensitivity to abrasion. It is likely that abrasion of gel beads is related to 'fatigue' of the gel materials.

    Application of artificially immobilized nitrifying cells is the topic of the last theme, in which the two formerly discussed themes are combined. In the work described in Chapter 6 the applicability of promising support materials was studied in more detail and a suitable support was used to immobilize nitrifying cells. High nitrification capacities could be reached in pretreated domestic wastewater, demonstrating that it is indeed possible to apply this system. The possibilities, drawbacks and prospects of immobilized cells for environmental applications are discussed further in Chapter 7. It is demonstrated there on the basis of the gathered knowledge that the immobilized-cell process has promising characteristics to meet the desired demands for future wastewater-treatment plants i.e. high biomass concentration, small volume of aeration tank, low liquid retention times, reduced sludge production and a high operational flexibility. Furthermore, it was calculated that the costs for production of immobilized cells are more than compensated for by the reduced costs for construction of a smaller reactor.

    Nitrification by immobilized cells
    Wijffels, R.H. - \ 1994
    Agricultural University. Promotor(en): J. Tramper. - S.l. : Wijffels - 188
    nitrificatie - cellen - immobilisatie - bradyrhizobiaceae - biotechnologie - chemische industrie - biochemie - geïmmobiliseerde cellen - nitrification - cells - immobilization - bradyrhizobiaceae - biotechnology - chemical industry - biochemistry - immobilized cells

    Nitrosomonas europaea and Nitrobacteragilis are nitrifying bacteria. Subsequently they oxidize ammonia to nitrite and nitrite to nitrate. Nitrification is a key process for the removal of nitrogen compounds from wastewater.

    Ammonia is removed from wastewater by supply of oxygen to a reactor containing nitrifying bacteria. The hydraulic retention time in a reactor with suspended nitrifying bacteria needs to be sufficient to prevent wash-out of the bacteria. By immobilization, i.e., attach or entrap the organisms in a support material and retain the support material in the reactor, it is possible to run a reactor under wash-out conditions and consequently execute the nitrification process in compact systems. In practice, biofilm reactors are most commonly used for this.

    In this research, artificial immobilized nitrifying bacteria are studied. The bacteria were suspended in a gel (carrageenan) and solid spheres were produced from this gel. The artificial immobilized cells were used as a model system for biofilm processes. By using the model system we were able to perform fundamental studies of growth within the support and substrate conversion by the immobilized cells.

    Growth of immobilized cells has been described qualitatively in chapter 2. Immobilized cells form small colonies as the result of growth. Initially, such expanding micro-colonies were observed over the entire gel bead, but after some time growth just under the surface of the spheres was faster because of diffusion limitation over the support.

    The qualitative observations have been quantified by modelling the significant transport processes (radial diffusion through the support and external mass transfer across the stagnant liquid layer surrounding the spheres), substrate consumption and growth (chapter 3). The model that we developed calculates substrate consumption and growth over time and shows for example that initially growth will take place all over the bead and after some time especially just under the surface of the beads. This is in agreement with our previous qualitative observations. The model has been validated experimentally by immobilized Nitrobacter agilis cells by determination of macroscopic substrate consumption rates, substrate profiles and biomass profiles (chapter 4 and 5). Experimental results were in close agreement with the model results.

    The model consisted of general equations and the parameters were obtained from separate experiments or literature. For this reason the model should be general applicable. To demonstrate the general applicability the model was also used for Nitrosomonas europaea (chapter 6). The calculated macroscopic consumption rates, however, were much larger and the biomass concentration much lower than we found experimentally. We could show that in this experiment not only diffusion limitation over gel beads but also diffusion limitation over the micro-colonies was important. The size of micro-colonies of Nitrosomonas europaea was much larger than of Nitrobacter agilis in the previous experiment. This has been the reason to implement growth of biomass by expansion of colonies and taking diffusion limitation over those colonies into account in our model. Validation of this colony-expansion model showed that experimental and simulated results agreed much better. Application of the colonyexpansion model to our previous experiment with Nitrobacter agilis also gave good results.

    The size of the micro-colonies formed is dependent to a higher degree on the amount of organisms that is immobilized than on the type of organism. At low initial biomass concentrations, a few but large colonies will be formed. At high initial biomass concentrations much more and much smaller colonies will be formed. Those smaller colonies experience less diffusion limitation and consequently the substrate-utilization capacity of such beads will be higher. This effect could be demonstrated, both experimentally and by simulations with immobilized Nitrosomonas europaea (chapter 7).

    The models that are presented in this thesis give insight in growth and substrateconversion capacity of immobilized cells. Our approach was general and as a consequence the insight that is obtained is not only important for nitrification, but can be used for other immobilized-cell processes as well.

    Engineering aspects of nitrification with immobilized cells
    Hunik, J.H. - \ 1993
    Agricultural University. Promotor(en): J. Tramper. - Wageningen : Hunik - 167
    chemische reacties - biotechnologie - chemische industrie - biochemie - cellen - immobilisatie - bradyrhizobiaceae - geïmmobiliseerde cellen - chemical reactions - biotechnology - chemical industry - biochemistry - cells - immobilization - bradyrhizobiaceae - immobilized cells

    Several aspects of a nitrification process with artificially immobilized cells in an airlift loop reactor have been investigated and are described in this thesis. In chapter 1 an overview of immobilization methods, suitable reactors, modelling, small-scale
    applications and scale-up strategy is given. The subjects of chapter 1 provide the starting point of the following chapters. Application of immobilized cells is beneficial for the nitrification process at high product and substrate concentrations and with a process temperature far below the optimal temperature of 30-35°C. In chapter 2 and 3 the kinetics of, respectively, Nitrosomonas europaea and Nitrobacter agilis cells at high product and substrate concentrations is presented. The results show a severe product inhibition of Nitrobacter agilis by nitrite, while Nitrosomonas europaea seems to be more sensitive
    for a high osmotic pressure. In chapter 4 a theoretical background of the immobilization method and further scale-up is presented. For the immobilization method the theory for the break-up of liquid jets, with Newtonian behaviour, is evaluated and a method to apply this theory for non-Newtonian liquids like a K-carrageenan solution is presented. In chapter 6 a dynamic model for the nitrification with immobilized Nitrosomonas europaea and Nitrobacter agilis cells is presented. The model includes mass-transfer rates, kinetic behaviour of the microorganisms, and reactor and gel bead properties. Predictions
    of reactor bulk concentrations of NH 4+, NO 2-and NO 3-(N-compounds) are given by the model together with concentration profiles of N-compounds, oxygen and biomass in the gel beads. A sensitivity analysis of the model parameters shows that the diffusion coefficient of oxygen in the gel beads and the radius of the gel beads are the most important parameters influencing the model output. The model is experimentally validated by means of reactor bulk concentrations and biomass profiles of Nitrosomonas europaea and Nitrobacter agilis in the gel beads. Predicted and measured values agree very well and the assumptions and equations used in the model seem to be valid. The biomass profiles of the two microorganisms co-immobilized in the gel beads are determined with immunofluorescence and a stereological method (chapter 5). The immunofluorescence technique was used to separate the Nitrosomonas europaea and Nitrobacter agilis colonies in the beads. From the position and diameter of the colonies it is possible to determine the spatial distribution of the two microorganisms in the gel beads. In chapter 7 the model is used as a tool to develop a strategy to scale-up the nitrification process with immobilized cells. A design for a large-scale application should be optimized with respect to the transport of oxygen to the immobilized cells in the gel beads. This is an advantage over the nitrification process with suspended cells, which is limited by the growth rate of the nitrifying bacteria. The growth of nitrifying bacteria is a slower process than the transport of oxygen to the cells. Some interesting aspects, which were not treated elsewhere in this thesis, are discussed in the general discussion in chapter 8.

    Plant cells : immobilization and oxygen transfer
    Hulst, A.C. - \ 1987
    Agricultural University. Promotor(en): J. Tramper, co-promotor(en): K. van 't Riet. - S.l. : Hulst - 121
    plantkunde - cellen - cytologie - immobilisatie - metabolisme - voeding - plantenfysiologie - celmetabolisme - celfysiologie - geïmmobiliseerde cellen - botany - cells - cytology - immobilization - metabolism - nutrition - plant physiology - cell metabolism - cell physiology - immobilized cells

    The study described in this thesis is part of the integrated project 'Biotechnological production of non-persistent bioinsecticides by means of plant cells invitro ' and was done in close cooperation with the research Institute Ital within the framework of NOVAPLANT. The plant cells used in this project were Tagetes species which produce thiophenes, naturally occurring biocides, particularly against nematodes.

    The objective of the study described in this thesis was to use immobilized plant cells or large plant cell aggregates for secondary metabolite production. In particular the upscaling of immobilization techniques for plant cells, the role of diffusion limitation of oxygen as a substrate on the immobilized plant cells and its effect on secondary metabolite production of the immobilized plant cells were subject of research.

    A literature survey on immobilized plant cells is presented in Chapter 2. The advantages of immobilized plant cells, several aspects concerning immobilization techniques, consequences of plant cell immobilization, immobilized plant cell reactors, and future prospects of immobilized plant cells are discussed in this Chapter.

    Chapter 3 deals with the application of the resonance nozzle as an immobilization technique with a high production capacity for plant cells as well as yeast cells in calcium alginate gel beads. It is found that this technique has a production capacity of two orders of magnitude larger than the conventional dripping technique with a needle. The viability of the cells after immobilization with the resonance nozzle was preserved. An extension of the applicability of the nozzle technique for thermogelling gel supports (k-carrageenan, agar and gellan gum) Is described in Chapter 4. Plant cells, yeast cells, bacterial cells and insect cells were used as model systems in the experiments.

    In Chapter 5, the occurrence of oxygen diffusion limitation of Daucuscarota cells in agarose, calcium alginate and (κ-carrageenan, is determined by respiration measurements of the immobilized cells in order to explain the enhanced pronounced secondary metabolite production with alginate immobilized plant cells from in the literature reported experiments. However, in our experiments no differences between the support materials could be observed.

    The effective diffusion coefficient for oxygen (ID e ) in the gel material is an important factor in mathematical model calculations in order to quantify the occurrence of oxygen diffusion limitation. Chapter 6 deals with the experiments in which (ID e ) was determined in different gel materials (calcium alginate, κ-carrageenan, gellan gum, agar and agarose) by measuring the oxygen diffusion from a well- stirred solution into gel beads, which were initially free of oxygen. A mathematical model was fitted on the experimental data resulting in the value of (ID e ) which was used In the following experiments.

    In several parts of this thesis a mathematical model was used for calculation of oxygen concentration profiles in gel beads containing plant cells or cell aggregates in order to visualize the occurrence of oxygen diffusion limitation. In Chapter 7 this model is tested on validity by experimental measurement of the oxygen concentration profiles in agarose beads containing respiring plant cells of Tagetesminuta . This was done with the aid of an oxygen microelectrode with a tip of 15 μm. The experimental and calculated oxygen concentration profiles correspond quite well.

    Chapter 8 deals with the effects of aggregate size and oxygen diffusion limitation on thiophene production and cell growth by cell aggregates of Tagetespatula . It is concluded that aggregate size is related to thiophene production by the observation of an 'optimum' aggregate size where the production is highest. Calculations of the oxygen concentration profiles showed that this could be due to the absence of oxygen In the centre of the aggregates.

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