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Koller M.,University of Graz | Dias M.M.S.,Institute Of La Vision | Rodriguez-Contreras A.,Polytechnic University of Catalonia | Kunaver M.,Slovenian National Institute of Chemistry | And 3 more authors.
Materials | Year: 2015

Liquefied wood (LW) prepared in a microwave process was applied as a novel; inexpensive precursor feedstock for incorporation of (R)-3-hydroxyvalerate (3HV) into polyhydroxyalkanoate (PHA) biopolyesters in order to improve the biopolyester's material quality; Cupriavidus necator was applied as microbial production strain. For proof of concept, pre-experiments were carried out on a shake flask scale using different mixtures of glucose and LW as carbon source. The results indicate that LW definitely acts as a 3HV precursor, but, at the same time, displays toxic effects on C. necator at concentrations exceeding 10 g/L. Based on these findings, PHA biosynthesis under controlled conditions was performed using a fed-batch feeding regime on a bioreactor scale. As major outcome, a poly(3HB-co-0.8%-3HV) copolyester was obtained displaying a desired high molar mass of Mw = 5.39 * 105 g/mol at low molar-mass dispersity (DM of 1.53), a degree of crystallinity (Xc) of 62.1%, and melting temperature Tm (176.3 °C) slightly lower than values reported for poly([R]-3-hydroxybutyrate) (PHB) homopolyester produced by C. necator; thus, the produced biopolyester is expected to be more suitable for polymer processing purposes. © 2015 by the authors.


Horvat P.,University of Zagreb | Koller M.,University of Graz | Braunegg G.,ARENA Association for Resource Efficient and Sustainable Technologies
World Journal of Microbiology and Biotechnology | Year: 2015

A review of the use of elementary flux modes (EFMs) and their applications in metabolic engineering covered with yield space analysis (YSA) is presented. EFMs are an invaluable tool in mathematical modeling of biochemical processes. They are described from their inception in 1994, followed by various improvements of their computation in later years. YSA constitutes another precious tool for metabolic network modeling, and is presented in details along with EFMs in this article. The application of these techniques is discussed for several case studies of metabolic network modeling provided in respective original articles. The article is concluded by some case studies in which the application of EFMs and YSA turned out to be most useful, such as the analysis of intracellular polyhydroxyalkanoate (PHA) formation and consumption in Cupriavidus necator, including the constraint-based description of the steady-state flux cone of the strain’s metabolic network, the profound analysis of a continuous five-stage bioreactor cascade for PHA production by C. necator using EFMs and, finally, the study of metabolic fluxes in the metabolic network of C. necator cultivated on glycerol. © 2015, Springer Science+Business Media Dordrecht.


Koller M.,University of Graz | Salerno A.,University of Graz | Muhr A.,University of Graz | Reiterer A.,University of Graz | Braunegg G.,ARENA Association for resource efficient and sustainable technologies
Materiali in Tehnologije | Year: 2013

Polyhydroxyalkanoates (PHAs) attract considerable attention as sustainable "green plastics" with a real potential to replace their petrol-based competitors in some applications in the not-too-distant future. To reach this goal PHAs must be able to compete with the established petrol-based plastics in both technical and economic terms. The current PHA production is based on prized substrates of high nutritional value such as sucrose, starch or vegetable oils. An alternative, carbon-rich industrial waste can be used as a suitable feedstock. This would contribute to making PHAs economically competitive and would avoid the conflict with human nutrition or animal feeding. Consequently, the decision about the location of the PHA-production facilities depends on the preferable in-house availability of such waste streams. The issue of competitive, large-scale PHA production in Europe was the topic of the ANIMPOL and the WHEYPOL projects. Both intended to develop novel processes for the transformation of abundant, locally available, renewable wastes. In the ANIMPOL case, waste lipids from slaughterhouses are converted to glycerol and a mixture of saturated and unsaturated fatty acid esters (FAEs), better known as biodiesel. The production of saturated FAEs is 50 000 t per year in Europe, decreasing the biodiesel performance as an engine fuel. However, within the project it was demonstrated that they can be efficiently metabolized to PHAs. In the WHEYPOL project attention was focused on 1.4 × 108 t per year of whey from dairies. This waste is of limited use and causes environmental concern. However, lactose, the main carbohydrate found in whey, can be used as a substrate in the WHEYPOL bioprocesses. These strategies demonstrate the feasibility of making "green plastics" competitive by integrating their manufacturing directly into the existing production lines, where the convertible waste streams accrue.


Novak M.,University of Zagreb | Koller M.,University of Graz | Braunegg G.,ARENA Association for Resource Efficient and Sustainable Technologies | Horvat P.,University of Zagreb
Chemical and Biochemical Engineering Quarterly | Year: 2015

The potential of poly(hydroxyalkanoates) (PHAs) to replace conventional plastic materials justifies the increasing attention they have drawn both at lab-scale and in industrial biotechnology. The improvement of large-scale productivity and biochemical/genetic properties of producing strains requires mathematical modeling and process/strain optimization procedures. Current models dealing with structurally diversified PHAs, both structured and unstructured, can be divided into formal kinetic, low-structured, dynamic, metabolic (high-structured), cybernetic, neural networks and hybrid models; these attempts are summarized in this review. Characteristic properties of specific groups of models are stressed in light of their benefit to the better understanding of PHA biosynthesis, and their applicability for enhanced productivity. Unfortunately, there is no single type of mathematical model that expresses exactly all the characteristics of producing strains and/or features of industrial-scale plants; in addition, the different requirements for modelling of PHA production by pure cultures or mixed microbial consortia have to be addressed. Therefore, it is crucial to sophisticatedly adapt and fine-tune the modelling approach accordingly to actual processes, as the case arises. For "standard microbial cultivations and everyday practices", formal kinetic models (for simple cases) and "low-structured" models will be appropriate and of great benefit. They are relatively simple and of low computational demand. To overcome the specific weaknesses of different established model types, some authors use hybrid models. Here, satisfying compromises can be achieved by combining mechanistic, cybernetic, and neural and computational fluid dynamics (CFD) models. Therefore, this hybrid modelling approach appears to constitute the most promising solution to generate a holistic picture of the entire PHA production process, encompassing all the benefits of the original modelling strategies. Complex growth media require a higher degree of model structuring. For scientific purposes and advanced development of industrial equipment in the future, real systems will be modelled by highly organized hybrid models. All solutions related to modelling PHA production are discussed in this review.


Koller M.,University of Graz | Salerno A.,University of Graz | Strohmeier K.,University of Graz | Schober S.,University of Graz | And 4 more authors.
Biocatalysis and Biotransformation | Year: 2014

Polyhydroxyalkanoates (PHAs) with tailored properties are needed to meet consumer demands regarding the use of eco-compatible biobased polymeric materials and relevant plastic items. Inserting 3-hydroxvalerate (3HV) monomeric units in PHA biopolyesters results in poly[(R)-3-hydroxybutyrate-co-(R)-3- hydroxyvalerate] (PHBHV) copolyesters aimed at their conversion into production of biodegradable eco-friendly plastic items. As inexpensive novel precursor substrate structurally related to 3HV, a mixture of odd-numbered carboxylic acids with 9-17 carbon atoms was produced by oxidative ozonolysis of alkenes. This mixture was successfully applied for biomediated PHBHV production by Cupriavidus necator. Applying this mixture as carbon substrate, a molar 3HV fraction exceeding 0.12 was obtained. The isolated copolyesters featured a low degree of crystallinity, narrow molar mass distribution, and low melting temperatures. These properties should make application of the novel 3HV-precursors interesting for large-scale production of easily processable copolyesters. © 2014 Informa UK, Ltd.


Koller M.,University of Graz | Chiellini E.,University of Pisa | Braunegg G.,ARENA Association for Resource Efficient and Sustainable Technologies
Chemical and Biochemical Engineering Quarterly | Year: 2015

The halobacterium Haloferax mediterranei was used to study the production of two types of biopolymers: The biopolyester poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) was accumulated as intracellular granules, whereas an extracelluar polysaccharide was excreted in parallel to biopolyester synthesis. After production, microbial re-use and degradation of these polymers under different conditions were investigated to assess the requirements for handling the product-rich fermentation broth prior to the downstream processing for product recovery. Degradation kinetics of the polymers and the impact of different storage conditions on molar mass of PHBV were studied. It turned out that the biotechnological fermentation process can be run without any sterility precautions. No major product losses were observed without pasteurization of fermentation broth after the stop of fermentation. In addition, neither PHBV nor EPS are re-utilized by the cells for biomass formation even if the culture is maintained under conditions of carbon starvation for an extended time.


Koller M.,University of Graz | Marsalek L.,University of Natural Resources and Life Sciences, Vienna | de Sousa Dias M.M.,French Institute of Health and Medical Research | Braunegg G.,ARENA Association for Resource Efficient and Sustainable Technologies
New Biotechnology | Year: 2016

Sustainable production of microbial polyhydroxyalkanoate (PHA) biopolyesters on a larger scale has to consider the ". four magic e": economic, ethical, environmental, and engineering aspects. Moreover, sustainability of PHA production can be quantified by modern tools of Life Cycle Assessment.Economic issues are to a large extent affected by the applied production mode, downstream processing, and, most of all, by the selection of carbon-rich raw materials as feedstocks for PHA production by safe and naturally occurring wild type microorganisms. In order to comply with ethics, such raw materials should be used which do not interfere with human nutrition and animal feed supply chains, and shall be convertible towards accessible carbon feedstocks by simple methods of upstream processing. Examples were identified in carbon-rich waste materials from various industrial braches closely connected to food production. Therefore, the article shines a light on hetero-, mixo-, and autotrophic PHA production based on various industrial residues from different branches. Emphasis is devoted to the integration of PHA-production based on selected raw materials into the holistic patterns of sustainability; this encompasses the choice of new, powerful microbial production strains, non-hazardous, environmentally benign methods for PHA recovery, and reutilization of waste streams from the PHA production process itself. © 2016 Elsevier B.V.


PubMed | University of Graz, Polytechnic University of Mozambique and ARENA Association for Resource Efficient and Sustainable Technologies
Type: Journal Article | Journal: New biotechnology | Year: 2015

Bacillus megaterium uyuni S29, isolated from the Bolivian salt lake Uyuni, displays a high capability to produce poly[(R)-3-hydroxybutyrate] (PHB) in industrial culture media. In order to analyze the influence of salt on biomass formation and PHB production, cultivations at different NaCl concentrations were carried out according to the salinity conditions of the habitats of the strains original isolation. In this preliminary report, the strain showed considerable adaptability to media of different salinity, obtaining the best results for both cellular growth and PHB production in media containing 45 g/L NaCl. The strain grew at 100 g/L NaCl and PHB production was observed even at high salt levels of 250 g/L without unwanted concurrent spore formation. Its tolerance to high salt concentrations together with auspicious PHB productivity makes this strain appealing not only for PHB production, but also for other biotechnological applications such as the treatment of salty wastewater; additional studies will be needed to further increase PHB productivity.


PubMed | University of Graz, University of Natural Resources and Life Sciences, Vienna, French Institute of Health and Medical Research and ARENA Association for Resource Efficient and Sustainable Technologies
Type: | Journal: New biotechnology | Year: 2016

Sustainable production of microbial polyhydroxyalkanoate (PHA) biopolyesters on a larger scale has to consider the four magic e: economic, ethical, environmental, and engineering aspects. Moreover, sustainability of PHA production can be quantified by modern tools of Life Cycle Assessment. Economic issues are to a large extent affected by the applied production mode, downstream processing, and, most of all, by the selection of carbon-rich raw materials as feedstocks for PHA production by safe and naturally occurring wild type microorganisms. In order to comply with ethics, such raw materials should be used which do not interfere with human nutrition and animal feed supply chains, and shall be convertible towards accessible carbon feedstocks by simple methods of upstream processing. Examples were identified in carbon-rich waste materials from various industrial braches closely connected to food production. Therefore, the article shines a light on hetero-, mixo-, and autotrophic PHA production based on various industrial residues from different branches. Emphasis is devoted to the integration of PHA-production based on selected raw materials into the holistic patterns of sustainability; this encompasses the choice of new, powerful microbial production strains, non-hazardous, environmentally benign methods for PHA recovery, and reutilization of waste streams from the PHA production process itself.

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