Andreas Schmid

Helmholtz-Zentrum für Umweltforschung, Leipzig, Saxony, Germany

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Publications (184)791.06 Total impact

  • Jan Volmer, Andreas Schmid, Bruno Bühler
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    ABSTRACT: Industrial bioprocess development is driven by profitability and eco-efficiency. It profits from an early stage definition of process and biocatalyst design objectives. Microbial bioprocess environments can be considered as synthetic technical microbial ecosystems. Natural systems follow Darwinian evolution principles aiming at survival and reproduction. Technical systems objectives are eco-efficiency, productivity, and profitable production. Deciphering technical microbial ecology reveals differences and similarities of natural and technical systems objectives, which are discussed in this review in view of biocatalyst and process design and engineering strategies. Strategies for handling opposing objectives of natural and technical systems and for exploiting and engineering natural properties of microorganisms for technical systems are reviewed based on examples. This illustrates the relevance of considering microbial ecology for bioprocess design and the potential for exploitation by synthetic biology strategies.
    Current Opinion in Microbiology 06/2015; 25:25-32. DOI:10.1016/j.mib.2015.02.002 · 7.22 Impact Factor
  • Karsten Lang, Katja Buehler, Andreas Schmid
    Advanced Synthesis & Catalysis 05/2015; 357(8). DOI:10.1002/adsc.201500205 · 5.54 Impact Factor
  • Christian David, Katja Bühler, Andreas Schmid
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    ABSTRACT: The application of segmented flow on a Synechocystis sp. PCC 6803 biofilm prevented excessive biomass formation and clogging by fundamentally changing the structure of the microbial community. It was possible to continuously operate a capillary microreactor for 5 weeks, before the experiment was actively terminated. The biofilm developed up to a thickness of 70-120 µm. Surprisingly, the biofilm stopped growing at this thickness and stayed constant without any detachment events occurring afterwards. The substrates CO2 and light were supplied in a counter-current fashion. Confocal microscopy revealed a throughout photosynthetically active biofilm, indicated by the red fluorescence of photo pigments. This control concept and biofilm reaction setup may enable continuous light driven synthesis of value added compounds in future.
    Journal of Industrial Microbiology 05/2015; 42(7). DOI:10.1007/s10295-015-1626-5 · 2.51 Impact Factor
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    ABSTRACT: For the investigation and comparison of microbial biofilms, a variety of analytical methods have been established, all focusing on different growth stages and application areas of biofilms. In this study, a novel quantitative assay for analysing biofilm maturation under the influence of continuous flow conditions was developed using the interesting biocatalyst Pseudomonas taiwanensis VLB120. In contrast to other tubular-based assay systems, this novel assay format delivers three readouts using a single setup in a total assay time of 40 h. It combines morphotype analysis of biofilm colonies with the direct quantification of biofilm biomass and pellicle formation on an air/liquid interphase. Applying the Tube-Assay, the impact of the second messenger cyclic diguanylate on biofilm formation of P. taiwanensis VLB120 was investigated. To this end, 41 deletions of genes encoding for protein homologues to diguanylate cyclase and phosphodiesterase were generated in the genome of P. taiwanensis VLB120. Subsequently, the biofilm formation of the resulting mutants was analysed using the Tube-Assay. In more than 60 % of the mutants, a significantly altered biofilm formation as compared to the parent strain was detected. Furthermore, the potential of the proposed Tube-Assay was validated by investigating the biofilms of several other bacterial species.
    Applied Microbiology and Biotechnology 05/2015; DOI:10.1007/s00253-015-6628-8 · 3.81 Impact Factor
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    ABSTRACT: Acidovorax sp. CHX100 has a remarkable ability for growth on short cycloalkanes (C5–C8) as a sole source of carbon and energy under aerobic conditions via an uncharacterized mechanism. Transposon mutagenesis of Acidovorax sp. CHX100 revealed a novel cytochrome P450 monooxygenase (CYP450chx) which catalyzed the transformation of cyclohexane to cyclohexanol. Primer walking methods categorized CYP450chx as cytochrome P450 class I taking into account its operon structure: monooxygenase, FAD oxidoreductase, and ferredoxin. CYP450chx was successfully cloned and expressed in Escherichia coli JM109. The activity of CYP450chx was demonstrated by means of the indole co-oxidation. Biotransformation capability of CYP450chx was confirmed through the catalysis of cycloalkanes (C5–C8) to their respective cyclic alcohols.
    Applied Microbiology and Biotechnology 05/2015; DOI:10.1007/s00253-015-6599-9 · 3.81 Impact Factor
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    ABSTRACT: The formation of stable emulsions in biphasic biotransformations catalyzed by microbial cells turned out to be a major hurdle for industrial implementation. Recently, a cost-effective and efficient downstream-processing approach, using supercritical carbon dioxide (scCO2 ) for both irreversible emulsion destabilization (enabling complete phase separation within minutes of emulsion treatment) and product purification via extraction has been proposed by Brandenbusch et al.(Biotechnology and Bioengineering 107:642-651, 2010). One of the key factors for a further development and scale-up of the approach is the understanding of the mechanism underlying scCO2 -assisted phase separation. A systematic approach was applied within this work to investigate the various factors influencing phase separation during scCO2 treatment (that is pressure, exposure of the cells to CO2 , and changes of cell surface properties). It was shown that cell toxification and cell disrupture are not responsible for emulsion destabilization. Proteins from the aqueous phase partially adsorb to cells present at the aqueous-organic interface, causing hydrophobic cell surface characteristics, and thus contribute to emulsion stabilization. By investigating the change in cell-surface hydrophobicity of these cells during CO2 treatment, it was found that a combination of catastrophic phase inversion and desorption of proteins from the cell surface is responsible for irreversible scCO2 mediated phase separation. These findings are essential for the definition of process windows for scCO2 -assisted phase separation in biphasic whole-cell biocatalysis. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
    Biotechnology and Bioengineering 05/2015; DOI:10.1002/bit.25655 · 4.16 Impact Factor
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    ABSTRACT: Emulsion stability plays a crucial role for mass transfer and downstream processing in organic-aqueous bioprocesses based on whole microbial cells. In this study, emulsion stability dynamics and the factors determining them during two-liquid phase biotransformation were investigated for stereoselective styrene epoxidation catalyzed by recombinant Escherichia coli. Upon organic phase addition, emulsion stability rapidly increased correlating with a loss of solubilized protein from the aqueous cultivation broth and the emergence of a hydrophobic cell fraction associated with the organic-aqueous interface. A novel phase inversion-based method was developed to isolate and analyze cellular material from the interface. In cell-free experiments, a similar loss of aqueous protein did not correlate with high emulsion stability, indicating that the observed particle-based emulsions arise from a convergence of factors related to cell density, protein adsorption, and bioreactor conditions. During styrene epoxidation, emulsion destabilization occurred correlating with product-induced cell toxification. For biphasic whole-cell biotransformations, this study indicates that control of aqueous protein concentrations and selective toxification of cells enables emulsion destabilization and emphasizes that biological factors and related dynamics must be considered in the design and modeling of respective upstream and especially downstream processes.
    Journal of Industrial Microbiology 04/2015; 42(7). DOI:10.1007/s10295-015-1621-x · 2.51 Impact Factor
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    ABSTRACT: Variability in whole-cell biocatalyst performance represents a critical aspect for stable and productive bioprocessing. In order to investigate whether and how oxygenase-catalyzed reactions are affected by such variability issues in solvent-tolerant Pseudomonas, different inducers, expression systems, and host strains were tested for the reproducibility of xylene and styrene monooxygenase catalyzed hydroxylation and epoxidation reactions, respectively. Significantly higher activity variations were found for biocatalysts based on solvent-tolerant Pseudomonas putida DOT-TIE and S12 compared with solvent-sensitive P. putida KT2440, Escherichia coli JM101, and solvent-tolerant Pseudomonas taiwanensis VLB120. Specific styrene epoxidation rates corresponded to cellular styrene monooxygenase contents. Detected variations in activity strictly depended on the type of regulatory system employed, being high with the alk- and low with the lac-system. These results show that the occurrence of clonal variability in recombinant gene expression in Pseudomonas depends on the combination of regulatory system and host strain, does not correlate with a general phenotype such as solvent tolerance, and must be evaluated case by case.
    Journal of Industrial Microbiology 04/2015; 42(6). DOI:10.1007/s10295-015-1615-8 · 2.51 Impact Factor
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    ABSTRACT: The recent progress in sustainable chemistry and in synthetic biology increased the interest of chemical and pharmaceutical industries to implement microbial processes for chemical synthesis. However, most organisms used in biotechnological applications are not evolved by Nature for the production of hydrophobic, non-charged, volatile, or toxic compounds. In order to overcome this discrepancy, bioprocess design should consist of an integrated approach addressing pathway, cellular, reaction, and process engineering. Highlighting selected examples, we show that surprisingly often Nature provides conceptual solutions to enable chemical synthesis. Complemented by established methods from (bio)chemical and metabolic engineering, these concepts offer potential strategies yet to be explored and translated into innovative technical solutions enabling sustainable microbial production of non-natural chemicals. Copyright © 2015 Elsevier Ltd. All rights reserved.
    Current Opinion in Biotechnology 03/2015; 35:52-62. DOI:10.1016/j.copbio.2015.03.010 · 8.04 Impact Factor
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    ABSTRACT: Relative and absolute quantification of proteins in biological and clinical samples are common approaches in proteomics. Until now, targeted protein quantification is mainly performed using a combination of HPLC-based peptide separation and selected reaction monitoring on triple quadrupole mass spectrometers. Here, we show for the first time the potential of absolute quantification using a direct infusion strategy combined with single ion monitoring (SIM) on a Q Exactive mass spectrometer. By using complex membrane fractions of Escherichia coli, we absolutely quantified the recombinant expressed heterologous human cytochrome P450 monooxygenase 3A4 (CYP3A4) comparing direct infusion-SIM with conventional HPLC-SIM. Direct-infusion SIM revealed only 14.7% (±4.1 (s.e.m.)) deviation on average, compared to HPLC-SIM and a decreased processing and analysis time of 4.5 min (that could be further decreased to 30 s) for a single sample in contrast to 65 min by the LC-MS method. Summarized, our simplified workflow using direct infusion-SIM provides a fast and robust method for quantification of proteins in complex protein mixtures.
    03/2015; 100. DOI:10.1016/j.euprot.2015.03.001
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    ABSTRACT: Metabolic engineering strategies mark a milestone for the fermentative production of bulk and fine chemicals. Yet, toxic products and volatile reaction intermediates with low solubilities remain challenging. Prominent examples are artificial multistep pathways like the production of perillyl acetate (POHAc) from glucose via limonene. For POHAc, these limitations can be overcome by mixed-culture fermentations. A limonene biosynthesis pathway and cytochrome P450 153A6 (CYP153A6) as regioselective hydroxylase are used in two distinct recombinant E. coli. POHAc formation from glucose in one recombinant cell was hindered by ineffective coupling of limonene synthesis and low rates of oxyfunctionalization. The optimization of P450 gene expression led to the formation of 6.20 ± 0.06 mg gcdw (-1) POHAc in a biphasic batch cultivation with glucose as sole carbon and energy source. Increasing the spatial proximity between limonene synthase and CYP153A6 by a genetic fusion of both enzymes changed the molar limonene/POHAc ratio from 3.2 to 1.6. Spatial separation of limonene biosynthesis from its oxyfunctionalization improved POHAc concentration 3.3-fold to 21.7 mg L(-1) as compared to a biphasic fermentation. Mixed-cultures of E. coli BL21 DE3 containing the limonene biosynthesis pathway and E. coli MG1655 harboring either CYP153A6, or alternatively a cymene monooxygenase, showed POHAc formation rates of 0.06 or 0.11 U gcdw (-1) , respectively. This concept provides a novel framework for fermentative syntheses involving toxic, volatile, or barely soluble compounds or pathway intermediates. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
    Biotechnology and Bioengineering 03/2015; DOI:10.1002/bit.25592 · 4.16 Impact Factor
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    ABSTRACT: Microscale cultivation systems are important tools to elucidate cellular dynamics beyond the population average and understand the functional architecture of single cells. However, there is scant knowledge about the bias of different microcultivation technologies itself on cellular functions. We therefore performed a systematic cross-platform comparison of three different microscale cultivation systems commonly harnessed in single-cell analysis: microfluidic non-contact cell traps driven by negative dielectrophoresis, microfluidic monolayer growth chambers, and semi-solid agarose pads. We assessed specific single cell growth rates, division rates and morphological characteristics of single Corynebacterium glutamicum cells and microcolonies as a bacterial model organism with medical and biotechnological relevance under standardized growth conditions. Strikingly, specific single-cell and microcolony growth rates µmax were robust and conserved for several cell generations with all three microcultivation technologies, whereas division rates cells grown on agarose pads deviated by up to 50% from cells cultivated in negative dielectrophoresis traps and monolayer growth chambers. Furthermore, morphological characteristics like cell lengths and division symmetries of individual cells were affected when grown on agarose pads. This indicated a significant impact of solid cultivation supports on cellular traits. The results demonstrate the impact of the microcultivation technology on microbial physiology for the first time and show the need of a careful selection and design of the microcultivation technology in order to allow unbiased analysis of cellular behavior.
    Lab on a Chip 02/2015; 15(8). DOI:10.1039/C4LC01270D · 5.75 Impact Factor
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    ABSTRACT: The efficient coupling of biotransformation steps to an existing fermentation pathway is an interesting strategy to expand the product portfolio of Corynebacterium glutamicum as whole-cell biocatalyst. This is especially challenging if the biotransformation step comprises a direct link to central metabolism, as in the case of α-ketoglutarate-dependent oxygenase catalysis. Aiming at trans-4-hydroxy-L-proline (Hyp) production from glucose in a minimal medium, the proline-4-hydroxylase gene from Dactylosporangium sp. strain RH1 was introduced into a proline-producing, isoleucine-bradytroph C. glutamicum strain. The production of proline was found to be induced by isoleucine limitation. Proline and Hyp production were found to depend differently on isoleucine limitation. Severe isoleucine limitation was shown to result in proline accumulation and low hydroxylation rates both in batch and continuous cultivation set-ups. The investigation of different steady states with various glucose:isoleucine molar ratios revealed that optimal conditions for Hyp production are met around a molar ratio of 46:1, where isoleucine limitation is sufficient to trigger proline production but the hydroxylation rate is high enough to convert the majority of formed proline to Hyp. A high cell-density fed-batch set-up was designed, capable of producing 7.1 g L−1 of Hyp from glucose in 23 h with 98.5% conversion of proline to Hyp. Reaction engineering, specifically the fine-tuning of the glucose:isoleucine concentration ratio, enabled control of the fermentation profile and thus the accumulation of the desired product Hyp from glucose in minimal and defined media. Biotechnol. Bioeng. © 2014 Wiley Periodicals, Inc.
    Biotechnology and Bioengineering 02/2015; 112(2). DOI:10.1002/bit.25442 · 4.16 Impact Factor
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    Christian Dusny, Andreas Schmid
    Microbial Biotechnology 01/2015; 8(1):23. DOI:10. 1111/1751-7915.12252 · 3.21 Impact Factor
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    ABSTRACT: Recent environmental economic developments generate a need for sustainable and cost-effective (microbial) processes for the production of high-volume, low-priced bulk chemicals. As an example, n-Butanol has, as a 2nd generation biofuel, beneficial characteristics compared to ethanol in liquid transportation fuel applications. The industrial revival of the classic n-butanol (ABE) fermentation requires process and strain engineering solutions for overcoming the main process limitations: product toxicity and low space-time yield (STY). Reaction intensification on the biocatalyst, fermentation, and bioprocess level can be based on economic and ecologic evaluations using quantifiable constraints. This review describes the means of process intensification for biotechnological processes. A quantitative approach is then used for the comparison of the massive literature on n-butanol fermentation. A comprehensive literature study – including key fermentation performance parameters – is presented and the results are visualized using the window of operation methodology. The comparison allowed the identification of the key constraints high cell densities, high strain stability, high specific production rate, cheap in situ product removal (ISPR), high n-butanol tolerance to operate ISPR efficiently, and cheap carbon source. It can thus be used as a guideline for the bioengineer during the combined biocatalyst, fermentation, and bioprocess development and intensification.This article is protected by copyright. All rights reserved
    Engineering in Life Sciences 01/2015; 15(1). DOI:10.1002/elsc.201400041 · 1.89 Impact Factor
  • Nadine Ladkau, Andreas Schmid, Bruno Bühler
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    ABSTRACT: Whole-cell biocatalysis has emerged as an important tool for the synthesis of value-added fine and bulk chemicals as well as pharmaceuticals. Especially, the rapid development of recombinant DNA technologies resulted in a shift from the exploitation of natural enzymes and pathways to the design of recombinant cell factories comprising heterologous enzymes and/or synthetic, orthologous pathways for the synthesis of industrially relevant compounds. This review discusses recent developments and concepts applied in the frame of multistep whole-cell biocatalysis along with representative examples.
    Current Opinion in Biotechnology 12/2014; 30:178–189. DOI:10.1016/j.copbio.2014.06.003 · 8.04 Impact Factor
  • Organic Process Research & Development 11/2014; 18(11):1516-1526. DOI:10.1021/op5002116 · 2.55 Impact Factor
  • Christian Dusny, Andreas Schmid
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    ABSTRACT: Life is based on the cell as the elementary replicative and self-sustaining biological unit. Each single cell constitutes an independent and highly dynamic system with a remarkable individuality in a multitude of physiological traits and responses to environmental fluctuations. However, with traditional population-based cultivation set-ups, it is not possible to decouple inherent stochastic processes and extracellular contributions to phenotypic individuality for two central reasons: the lack of environmental control and the occlusion of single-cell dynamics by the population average. With microfluidic single-cell analysis as a new cell assay format, these issues can now be addressed, enabling cultivation and time-resolved analysis of single cells in precisely manipulable extracellular environments beyond the bulk. In this article, we explore the interplay of cellular physiology and environment at a single-cell level. We review biological basics that govern the functional state of the cell and put them in context with physical fundamentals that shape the extracellular environment. Furthermore, the significance of single-cell growth rates as pivotal descriptors for global cellular physiology is discussed and highlighted by selected studies. These examples illustrate the unique opportunities of microfluidic single-cell cultivation in combination with growth rate analysis, addressing questions of fundamental bio(techno)logical interest.
    Environmental Microbiology 10/2014; 17(6). DOI:10.1111/1462-2920.12667 · 6.24 Impact Factor
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    ABSTRACT: Catalytic biofilms minimize reactant toxicity and maximize biocatalyst stability in selective transformations of chemicals to value-added products in continuous processes. The scaling up of such catalytic biofilm processes is challenging, due to fluidic and biological parameters affording a special reactor design affecting process performance. A solid support membrane aerated biofilm reactor was optimized and scaled-up to yield gram amounts of (S)-styrene oxide, a toxic and instable high value chemical synthon. A sintered stainless steel membrane unit was identified as an optimal choice as biofilm substratum and for high oxygen mass transfer. A stable expanded PTFE (ePTFE) membrane was best suited for in situ substrate delivery and product extraction. For the verification of scalability, catalytic biofilms of Pseudomonas sp. strain VLB120ΔC produced (S)-styrene oxide to an average concentration of 390 mM in the organic phase per day (equivalent to 24.4 g Laq (-1) day(-1) ). This productivity was gained by efficiently using the catalyst with an excellent product yield on biomass of 13.6 gproduct gbiomass (-1) . This product yield on biomass is in the order of magnitude reported for other continuous systems based on artificially immobilized biocatalysts and is fulfilling the minimum requirements for industrial biocatalytic processes. Overall, 46 g of (S)-styrene oxide were produced and isolated (purity: 99%; ee: > 99.8%. yield: 30%). The productivity is in a similar range as in comparable small scale biofilm reactors highlighting the large potential of this methodology for continuous bioprocessing of bulk chemicals and biofuels.
    Biotechnology Journal 10/2014; 9(10). DOI:10.1002/biot.201400341 · 3.71 Impact Factor
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    ABSTRACT: Biofilm reactors are often mass transfer limited due to excessive biofilm growth, impeding reactor performance. Fluidic conditions play a key role for biofilm structural development and subsequently for overall reactor performance. Continuous interfacial forces generated by aqueous-air segmented flow are controlling biofilm structure and diminish mass transfer limitations in biofilm microreactors. A simple three step method allows the formation of robust biofilms under aqueous-air segmented flow conditions: a first-generation biofilm is developing during single phase flow, followed by the introduction of air segments discarding most of the established biofilm. Finally, a second-generation, mature biofilm is formed in the presence of aqueous-air segments. Confocal laser scanning microscopy experiments revealed that the segmented flow supports the development of a robust biofilm. This mature biofilm is characterized by a three to four fold increase in growth rate, calculated from an increase in thickness, a faster spatial distribution (95% surface coverage in 24h), and a significantly more compact structure (roughness coefficient <1), as compared to biofilms grown under single phase flow conditions. The applicability of the concept in a segmented flow biofilm microreactor was demonstrated using the epoxidation of styrene to (S)-styrene oxide (ee > 99.8%) catalyzed by Pseudomonas sp. strain VLB120▵C cells in the mono-species biofilm. The limiting factor affecting reactor performance was oxygen transfer as the volumetric productivity rose from 11 to 46 g Ltube−1 day−1 after increasing the air flow rate. In summary, different interfacial forces can be applied for separating cell attachment and adaptation resulting in the development of a robust catalytic biofilm in continuous microreactors. Biotechnol. Bioeng. © 2014 Wiley Periodicals, Inc.
    Biotechnology and Bioengineering 09/2014; 111(9). DOI:10.1002/bit.25256 · 4.16 Impact Factor

Publication Stats

5k Citations
791.06 Total Impact Points


  • 2014–2015
    • Helmholtz-Zentrum für Umweltforschung
      Leipzig, Saxony, Germany
    • Universität Stuttgart
      • Institute for Biochemical Engineering
      Stuttgart, Baden-Württemberg, Germany
  • 2005–2015
    • Technische Universität Dortmund
      • • Laboratory of Chemical Biotechnology (BT)
      • • Chair of Chemical Biology
      Dortmund, North Rhine-Westphalia, Germany
  • 2010–2011
    • Leibniz-Institut für Analytische Wissenschaften
      Dortmund, North Rhine-Westphalia, Germany
  • 2008
    • Max Planck Institute of Molecular Physiology
      Dortmund, North Rhine-Westphalia, Germany
    • Ewha Womans University
      Sŏul, Seoul, South Korea
  • 2006
    • Delft University of Technology
      • Department of Biotechnology
      Delft, South Holland, Netherlands
  • 1999–2006
    • Eawag: Das Wasserforschungs-Institut des ETH-Bereichs
      Duebendorf, Zurich, Switzerland
  • 2004
    • Hochschule für Technik Zürich
      Zürich, Zurich, Switzerland
  • 2002
    • Wageningen University
      • Laboratory of Biochemistry
      Wageningen, Gelderland, Netherlands
  • 1998–2002
    • ETH Zurich
      • Institute of Energy Technology
      Zürich, ZH, Switzerland